Compounds for control of appetite

ABSTRACT

This invention relates generally to neuropeptide Y (“NPY”) Y 4  receptor agonists including pancreatic polypeptide (PP), analogs thereof, and peptide fragments of PP, e.g. PP(32-36), and analogs thereof, to pharmaceutical compositions containing such Y 4  receptor agonists, and to methods for treatment of mammals using the same. The NPY Y 4  receptor agonists may be administered to mammals either alone or in combination with NPY Y 2  receptor agonists including peptide YY (PYY) (3-36), analogs thereof, and to peptide fragments of PYY(3-36), e.g. PYY(22-36) and PYY(25-36), and analogs thereof, such as to control food intake in mammals, blood pressure, cardiovascular response, libido, circadian rhythm, hyperlipidimia, chronic pancreatitis, and nonalcoholic fatty liver disease including nonalcoholic steatohepatitis.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH

The U.S. Government has a paid-up license in this invention and theright in limited circumstances to require the patent owner to licenseothers on reasonable terms as provided for by the terms ofGrant/Contract No. GM47122-08S1 awarded by the National Institutes ofHealth.

FIELD OF THE INVENTION

This invention relates generally to neuropeptide Y (“NPY”) Y₄ receptoragonists including pancreatic polypeptide (PP), analogs thereof, and topeptide fragments of PP, and analogs thereof, and to methods fortreatment of mammals using the same and, more specifically, theinvention is directed to NPY Y₄ receptor agonists including PP(32-36),and analogs thereof, to pharmaceutical compositions containing suchpentapeptides, and to methods of treatment of mammals using suchpentapeptides. The NPY Y₄ receptor agonists may be administered tomammals either alone or in combination with NPY Y₂ receptor agonistsincluding peptide YY (PYY) (3-36), analogs thereof, and to peptidefragments of PYY(3-36), and analogs thereof, such as to control foodintake in mammals.

BACKGROUND OF THE INVENTION

Obesity is a major disorder affecting as much as one third of the NorthAmerican population. Several studies have shown that such individualsare at increased risk in developing cardiovascular disease (hypertensionand hyper-cholesterolemia), diabetes and several types of cancer. Theeffective treatment of obesity, however, remains a largely unachievedgoal. Existing pharmaco-therapeutic approaches to weight loss involvethe use of amphetamine-based agents such as amphetamine, diethylpropion,mazindol and fenfluramine which act directly on the central nervoussystem (“CNS”) to lower food intake by modulating dopaminergic,adrenergic and/or serotonergic mechanisms. Although weight loss can beachieved with such agents, their use is restricted due to CNSside-effects, potential addiction liability and the production oftolerance to their actions, with chronic administration leading topotential depression, vestibular disturbances, hallucinations andaddiction, as well as interference with the actions of other drugs, suchas MAO inhibitors and antihypertensives. There is also a subpopulationof obese patients that is refractory to present anorectic drugtreatments. The medical need is high for an effective anorectic agentthat overcomes the above disadvantages of existing therapies. Ofparticular need are agents which act by alternative mechanisms tomodulate food intake and/or metabolism.

Throughout this application, various publications are referenced. Thedisclosure of these publications is hereby incorporated by referenceinto this application to describe more fully the art to which thisinvention pertains.

Neuropeptide Y (“NPY”)

Neuropeptides are small peptides originating from large precursorproteins synthesized by peptidergic neurons and endocrine/paracrinecells. They hold promise for treatment of neurological, psychiatric, andendocrine disorders (De Wied, D. In: Neuropeptides: Basics andPerspectives (Elsevier, Amsterdam-New York-Oxford), 1990). Often theprecursors contain multiple biologically active peptides. There is greatdiversity of neuropeptides in the brain caused by alternative splicingof primary gene transcripts and differential precursor processing. Theneuropeptide receptors serve to discriminate between ligands and toactivate the appropriate signals. Thus, it is expected that thereceptors for neuropeptides consist of a large number of members.

Neuropeptide Y (NPY), a 36-amino acid peptide, is the most abundantneuropeptide to be identified in mammalian brain. Human NPY has theformula:H-Tyr-Pro-Ser-Lys-Pro-Asp-Asn-Pro-Gly-Glu-Asp-Ala-Pro-Ala-Glu-Asp-Met-Ala-Arg-Tyr-Tyr-Ser-Ala-Leu-Arg-His-Tyr-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH2(SEQ. ID. NO. 1). Porcine and rat NPY have the same sequence except forLeu instead of Met in the 17-position.

NPY is an important regulator in both the central and peripheral nervoussystems (Heilig, M. and E. Widerlov. Neuropeptide Y: an overview ofcentral distribution, functional aspects, and possible involvement ofneuropsychiatric illnesses. Acta Psychiatr. Scand. 82:95-114 (1990)) andinfluences a diverse range of physiological parameters, includingeffects on psychomotor activity, food intake, central endocrinesecretion, and vasoactivity in the cardiovascular system. Highconcentrations of NPY are found in the sympathetic nerves supplying thecoronary, cerebral, and renal vasculature and has contributed tovasoconstriction. NPY binding sites have been identified in a variety oftissues, including spleen (Lundberg, J. M., A. Hemsen, O. Larsson, A.Rudehill, A. Saria, and B. Fredholm. Neuropeptide Y receptor in pigspleen: binding characteristics, reduction of cyclic AMP formation andcalcium antagonist inhibition of vasoconstriction. Eur. J. Pharmacol.145:21-29 (1988)), intestinal membranes, brain (Hinson, J., C. Rauh, andJ. Coupet. Neuropeptide Y stimulates inositol phospholipid hydrolysis inrat brain microprisms. Brain Response. 446:379-382 (1988)), aorticsmooth muscle (Mihara, S., Y. Shigeri, and M. Fujimoto. NeuropeptideY-induced intracellular Ca2+ increase in vascular smooth muscle cells.FEBS Lett. 259: 79-82 (1989)), kidney, testis, and placenta (Dumont, Y.,J. C. Martel, A. Fournier, S. St.-Pierre, and R. Quiron. Neuropeptide Yand neuropeptide Y receptor subtypes in brain and peripheral tissues.Prog. Neurobiol. 38:125-167 (1992)). In addition, binding sites havebeen reported in a number of rat and human cell lines (e.g. Y1 inSK-N-MC, MC-IXC, CHP-212, and PC12 cells; Y2 in SK-N—Be(2), CHP-234, andSMS-MSN)(Grundemar, L., S. P. Sheikh, and C. Wahlestedt, In: The Biologyof Neuropeptide Y and Related Peptides. (Humana Press, Inc., Totawa,N.J.), (1992)).

NPY forms a family (called the pancreatic polypeptide family) togetherwith pancreatic polypeptide (PP) and peptide YY (PYY) which all consistof 36 amino acids and have a common tertiary structure, the so-calledPP-fold (Glover, I. D., D. J. Barlow, J. E. Pitts, S. P. Wood, I. J.Tickle, T. L. Blundell, K. Tatemoto, J. R. Kimmel, A. Wollmer, W.Strassburger, and Y.-S. Zhang. Conformational studies of the pancreaticpolypeptide hormone family. Eur. J. Biochem. 142:379-385 (1985)).Specific features of this family include a polyproline helix in residues1 through 8, beta-turn in residues 9 through 14, an alpha-helix inresidues 15 through 30, an outward-projecting C-terminus in residues 30through 36, and a carboxy terminal amide (Schwartz, T. W., J.Fuhlendorff, L. L. Kjems, M. S. Kristensen, M. Vervelde, M. O'Hare, J.L. Krstenansky, and B. Bjornholm. Signal epitopes in thethree-dimensional structure of neuropeptide Y. Ann. N.Y. Acad. Sci.611:35-47 (1990)). The C-terminal amidated residue of these peptidesappears to be essential for biological activity (Wahlestedt et al.,1986). Studies with peptide fragments of NPY have indicated thatmultiple NPY receptor subtypes exist (Wahlestedt, C., N. Yanaihara, andR. Hakanson. Evidence for different pre- and postjunctional receptorsfor neuropeptide Y and related peptides. Regul. Pept. 13:307-318(1986)). Specifically, six receptor subtypes, denoted as Y1, Y2, Y3, Y4,Y5, and Y6, are understood to mediate the actions of NPY with eachto-date, except for Y3, having been cloned.

The Y1, Y2, Y4, and Y5 receptors have been proposed to regulate feedingbehavior, i.e. food intake, in subjects. A key pharmacological featurewhich distinguishes Y1 from Y2 is the fact that the Y1 receptor (and notthe Y2 receptor) responds to an analog of NPY modified at residues 31and 34 ([Leu31,Pro34]NPY), whereas the Y2 receptor (and not the Y1receptor) has high affinity for the NPY peptide carboxyl-terminalfragment NPY-(13-36)(Fuhlendorff, J., U. Gether, L. Aakerlund, N.Langeland-Johansen, H. Thogersen, S. G. Melberg, U. B. Olsen, O.Thastrup, and T. W. Schwartz. [Leu31,Pro34]Neuropeptide Y: A specific Y1receptor agonist. Proc. Natl. Acad. Sci. USA 87:182-186 (1990)). NPYanalogs and N-terminally-shortened fragments, e.g. NPY(18-36), whichcontain one or more specific D-isomer substitutions for the naturallyoccurring residues (as well as pharmaceutically acceptable saltsthereof), dispersed in a pharmaceutically acceptable liquid or solidcarrier, can be administered to mammals, including humans, tosubstantially lower blood pressure over an extended period of time or tocounteract hypertension.

Experimental and clinical observations also have supported the conceptthat neuropeptides play central roles in neurotransmission as well asthe regulation of secretory functions of adenohypophysial, pancreatic,adrenalcortical and gut cells. Among the thirty or so neuropeptides thathave been implicated in neuronal function in the mammalian centralnervous system, several have also been suggested to function asneurotransmitters or neuromodulators primarily in afferent neurons.

An additional action of NPY is to decrease cardiac contractility(inotropy). This is an extremely important action of NPY, because it isknown that, under many circumstances in which inotropy is decreased,diseases of life-threatening importance, e.g. congestive heart failureand cardiogenic shock, are associated with probable increased release ofNPY into the blood. Prevention of NPY release, using a presynaptic NPYagonist, or NPY's action, using a postsynaptic NPY antagonist, may bebeneficial in these disease states.

NPY has also been reported to produce coronary artery vasoconstrictionand thereby may decrease myocardial blood flow resulting in myocardialischemia. Such a circumstance can result in angina pectoris or, undermore severe circumstances, may result in myocardial infarction anddeath. In recent years, several classes of drugs have proven effectivein dilating coronary arteries to prevent such events.

Peptide YY (“PYY”)

Peptide YY (PYY) is a 36-residue peptide amide isolated originally fromporcine intestine, and localized in the endocrine cells of thegastrointestinal tract and pancreas (Tatemoto et al. Proc. Natl. Acad.Sci. 79:2514, 1982). Peptide YY has N-terminal and C-terminal tyrosineamides; accordingly, these two tyrosines give PYY its name (Y representsthe amino acid tyrosine in peptide nomenclature). In addition, PYYshares a number of central and peripheral regulatory roles with itshomologous peptide Neuropeptide Y (NPY), which was originally isolatedfrom porcine brain (Tatemoto, Proc. Natl. Acad. Sci. 79:5485, 1982). PYYis localized in intestinal cells; NPY, in contrast, is present in thesubmucous and myenteric neurons that innervate the mucosal and smoothmuscle layers, respectively (Ekblad et al. Neuroscience 20:169, 1987).Both PYY and NPY are believed to inhibit gut motility and blood flow(Laburthe, Trends Endocrinol. Metab. 1:168, 1990), and they are alsothought to attenuate basal (Cox et al. Br. J. Pharmacol. 101:247, 1990;Cox et al. J. Physiol. 398:65, 1988; Cox et al. Peptides 12:323, 1991;Friel et al. Br. J. Pharmacol. 88:425, 1986) and secretatogue-inducedintestinal secretion in rats (Lundberg et al. Proc. Natl. Acad. Sci USA79:4471, 1982; Playford et al. Lancet 335: 1555, 1990) and humans(Playford et al., supra), as well as stimulate net absorption (MacFadyenet al. Neuropeptides 7:219, 1986). Elevated plasma PYY levels have beenreported in individuals suffering from several conditions that causediarrhea (Adrian et al. Gastroenterology 89:1070, 1985). Taken together,these observations suggest that PYY and NPY are released into thecirculation after a meal (Adrian et al. Gastroenterology 89:1070, 1985:Balasubramaniam et al. Neuropeptides 14:209, 1989), and, thus may play aphysiological role in regulating intestinal secretion and absorption,serving as natural inhibitors of diarrhea.

A high affinity PYY receptor system that exhibits a slightly higheraffinity for PYY than NPY has been characterized in rat intestinalepithelia (Laburthe et al. Endocrinology 118:1910, 1986; Laburthe,Trends Endocrinol. Metabl. supra) and shown to be negatively coupled toadenylate cyclase (Servin et al. Endocrinology 124:692, 1989).Consistently, PYY exhibited greater antisecretory potency than NPY involtage clamped preparations of rat small intestine (Cox et al. J.Physiol. supra), while C-terminal fragments of NPY were found to be lesseffective in their antisecretory potency than PYY (Cox et al. Br. J.Pharmacol, supra). Structure-activity studies using several partialsequences have led to the identification of PYY(22-36) as the activesite for interacting with intestinal PYY receptors (Balasumbramaniam etal. Pept. Res. 1:32, 1988). This intestinal PYY-preferring receptor hasnow been cloned and shown to be identical to the Y₂ receptors clonedfrom the brain (Goumain et al. Mol Pharmacol 60:124-134, 2001).

In addition, PYY has been implicated in a number of physiologicalactivities including nutrient uptake (see, e.g., Bilcheik et al.Digestive Disease Week 506:623, 1993), cell proliferation (see, e.g.,Laburthe, Trends Endocrinol. Metab. 1:168, 1990; Voisin et al. J. Bio.Chem, 1993), lipolysis (see, e.g., Valet et al. J. Clin. Invest. 85:291,1990), and vasoconstriction (see, e.g., Lundberg et al., Proc. Natl.Acad. Sci, USA 79:4471, 1982).

The amino acid sequences of porcine and human PYY are as follows:porcine PYY: YPAKPEAPGEDASPEELSRYYASLRHYLNLVTR (SEQ. ID. NO. 2) QRY,human PYY: YPIKPEAPGEDASPEELNRYYASLRHYLNLVTR (SEQ. ID. NO. 3) QRY.The amino acid sequences for dog PYY and for rat PYY are the same asthat of porcine PYY.

With respect to PYY, it has been reported previously that peripheraladministration of PYY(3-36), a NPY Y₂-preferring ligand, can onperipheral administration attenuate food intake in normal and fastedmice and rats as well as in normal and obese humans (Nature 418:650-654;2002, N Engl J Med 349:941-948; 2003). The anorexigenic actions ofPYY(3-36) are suggested to be mediated by arcuate nucleus Y₂ receptors.One advantage of using Y2 selective ligands is that they can suppressthe food intake on peripheral administration, whereas Y1 and Y5selective ligands, as described above, have to penetrate the BBB tomodulate food intake.

In addition to interacting with the Y2 receptor, PYY(3-36) can potentlyactivate Y4 and Y5 receptors. Consequently, the inventor previouslydeveloped Y2 receptor selective agonists that are based on PYY(22-36)and PYY(25-36) (See U.S. Pat. Nos. 5,604,203, and 6,046,167 toBalasubramaniam) which are devoid of activities at the other NPYreceptors including Y1, Y4, and Y5 at concentrations up to 20,000 nM.Preferred PYY(25-36) analogs include N-α-Ac-[Trp³⁰]PYY(25-36)-NH₂ (SEQ.ID. NO. 4), referred to as BWX-115, N-α-Ac-[Trp²⁷,ψ^(35/36)]PYY(25-36)-NH₂ (SEQ. ID. NO. 5), referred to as BT-56, andN-α-Ac-[Trp³⁰, ψ^(35/36)]PYY(25-36)-NH₂ (SEQ. ID. NO. 6), referred to asBT-123, and the PYY(22-36) analog N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹,ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO. 7), referred to as B-48, [whereinψ in the foregoing formulas is —CH2-NH—] which can be used to controlfood intake in animals and humans.

Pancreatic Peptide (“PP”)

Pancreatic peptide (PP) is a 36-amino-acid secretory peptide that ispredominantly produced by the pancreas and released in response tonutrient stimuli.

The amino acid sequence of human PP is as follows:APLEPVYPGDNATPEQMAQYAADLRRYINMLTR (SEQ. ID. NO. 8) PRY.

PP, like PYY, shares a number of central and peripheral regulatory roleswith its homologous peptide NPY. It has been reported recently that PPcan attenuate body weight increase in mice through inhibiting foodintake and increasing energy expenditures (Gastroenterology124:1325-1336; 2003). These actions are mediated by altering theexpression of orexigenic (NPY, orexin & ghrelin downregulated) andanorexigenic (urocortin upregulated) peptides, and decreasing gastricemptying and activity of the vagovagal or vago-sympathetic reflex arc.In addition, the anorectic effects of PP have been suggested to bemediated by the Y₄ receptors in the area postrema (AP) because: a)autoradiography following peripheral administration of PP revealed highaccumulation of PP in AP; b) expression of c-fos is seen in AP followingiv administration of PP; and c) high density of Y₄ receptors are presentin AP (Brain Res 760:137-149; 1997).

One advantage of using Y4 selective ligands, like the Y2 selectiveligand, is that they can suppress the food intake on peripheraladministration, whereas Y1 and Y5 selective ligands, as described above,have to penetrate the BBB to modulate food intake. Native PP may not bean ideal candidate for suppressing food intake because of its shorthalf-life of about six minutes and its interactions with Y5 receptors.

Accordingly, it would be desirable to develop NPY Y₄ receptor agoniststhat can be used alone or mixed with NPY Y₂ receptor agonists, asdisclosed in U.S. Pat. Nos. 5,604,203, and 6,046,167 to Balasubramaniam,which are herein incorporated by reference, for controlling an NPYmediated physiological response in a subject, such as to control foodintake in mammals, and that are expected to prove useful in thetreatment of weight problems (e.g. obesity and diabetes), eatingdisorders, and such.

SUMMARY OF THE INVENTION

This invention is directed to neuropeptide Y (“NPY”) Y₄ receptoragonists including pancreatic polypeptide (PP), analogs thereof, andpeptide fragments of PP, e.g. PP(32-36), and analogs thereof, topharmaceutical compositions containing such Y₄ receptor agonists, and tomethods for treatment of mammals using the same. The NPY Y₄ receptoragonists may be administered to mammals either alone or in combinationwith NPY Y₂ receptor agonists including peptide YY (PYY) (3-36), analogsthereof, and to peptide fragments of PYY(3-36), e.g. PYY(22-36) andPYY(25-36), and analogs thereof, as disclosed in U.S. Pat. Nos.5,604,203, and 6,046,167 to Balasubramaniam, which are hereinincorporated by reference, such as to control food intake in mammals,blood pressure, cardiovascular response, libido, circadian rhythm,hyperlipidimia, chronic pancreatitis, and nonalcoholic fatty liverdisease including nonalcoholic steatohepatitis.

Accordingly, in one aspect, the present invention features NPY Y₄receptor agonists of PP(32-36) and analogs thereof, such pentapeptideanalogs having the formula:

wherein:

-   each X, independently, is an aromatic amino acid;-   each Y, independently, is an amino acid having a guanidino group;-   each Z, independently, is an aliphatic amino acid;-   n=1, 2, 3 or 4; and-   wherein the compound optionally includes one or two pseudopeptide    bonds where each pseudopeptide bond is independently selected from    —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.

A preferred compound of formula (I) includes,

(which is referred to as B-74).

In another aspect, the present invention features pentapeptide analogsof PP(32-36) having the formula:

wherein:

-   each X, independently, is an aromatic amino acid;-   each Y, independently, is an amino acid having a guanidino group;-   each Z, independently, is an aliphatic amino acid;-   A1 and A2, independently, are selected from Cys, Pen, Glu, Asp, Lys,    and Dpr; and-   wherein the compound optionally includes one or two pseudopeptide    bonds where each pseudopeptide bond is independently selected from    —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.

A preferred compound of formula (II) includes,

In another aspect, the present invention features pentapeptide analogsof PP(32-36) having the formula:H—[X—Y-Z-Y—X]_(n)—NH₂  (III)

wherein:

-   each X, independently, is an aromatic amino acid;-   each Y, independently, is an amino acid having a guanidino group;-   each Z, independently, is an aliphatic amino acid;-   n=1, 2, 3 or 4; and-   wherein the compound optionally includes one or two pseudopeptide    bonds where each pseudopeptide bond is independently selected from    —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.

Preferred compounds of formula (III) includeH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ. ID. NO. 9) orH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ.ID. NO. 10).

In another aspect, NPY Y₄ receptor agonists, such as those of formulas(I-III), are mixed or combined with NPY Y₂ receptor agonists, such asanalogs of PYY(25-36) or PYY(22-36), for controlling an NPY mediatedphysiological response in a subject, e.g. to suppress appetite, suchanalogs of PYY(25-36) and of PYY(22-36), respectively, having formulas(IV) and (V) below:

wherein:

-   the N-terminal amino acid is bonded to R¹ and R²;-   Y is a chain of 0-4 amino acids, inclusive, where the C-terminal    amino acid has a carboxylamide group; which is independently bonded    to R³ and R⁴, e.g.,-   R¹ and R² are each independently bonded to the amino group of the    N-terminal amino acid and selected from H, (C₁-C₁₂)alkyl,    (C₆-C₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl;-   R³ and R⁴ are each independently bonded to the amide group of the    C-terminus amino acid, e.g.,    -   (where R denotes the side chain group of the amino acid, e.g.        R═H in Gly, etc.), and selected from H, (C₁-C₁₂)alkyl (e.g.        methyl), (C₆-C₁₈)aryl (e.g. phenyl), (C₁-C₁₂)acyl (formyl,        acetyl, napthaleneacetyl, and myristoyl), (C₇-C₁₈) aralkyl (e.g.        benzyl), and (C₇-C₁₈)alkaryl (e.g. p-methlyphenyl);-   A²⁵ is Arg, Lys, homo-Arg, diethyl-homo-Arg, lys-ε-NH—R where R is    H, a branched or straight chain (C₁-C₁₀) alkyl group, or an aryl    group, Orn or is deleted;-   A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His, β-pyrozolylalanin,    N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-e-NH—R where R    is H, a branched or straight chain (C₁-C₁₀) alkyl group, or an aryl    group, Orn or is deleted;-   A²⁷ is an aromatic amino acid:-   A²⁸ is Leu, Ile, Val, Trp Nle, Nva, Aib, Anb, or N-Me-Leu;-   A²⁹ is Asn, Ala, Gin, Fly, Trp, or N-Me-Asn;-   A³⁰ is Leu, Ile, Val, Trp, Nle, Nva, Aib, Anb, or N-Me-Leu;-   A³¹ is Val, Ile, Trp, Nva, Aib, Anb, or N-Me-Val;-   A³² is Thr, Ser, N-Me-Ser, N-Me-Thr, or D-Trp; and-   wherein the compound optionally includes one or two pseudopeptide    bonds where each pseudopeptide bond is independently selected from    —CH₂—NH—, —CH₂—S—, —CH₂CH₂—, —CH₂—O— and CH₂—CO—;    wherein-   X is a chain of 0-5 amino acids, inclusive, where the N-terminal    amino acid is bonded to R¹ and R² by the nitrogen of the amino group    of the N-terminal amino acid;-   Y is a chain of 0-4 amino acids, inclusive, where the C-terminal    amino acid has a carboxylamide group; which is independently bonded    to R³ and R⁴, e.g.,-   R¹ and R² are each independently bonded to the amino group of the    N-terminal amino acid and selected from H, (C₁-C₁₂)alkyl,    (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl;-   R³ and R⁴ are each independently bonded to the amide group of the    C-terminus amino acid, e.g.,    -   (where R denotes the side chain group of the amino acid, e.g.        R═H in Gly, etc.), and selected from H, (C₁-C₁₂)alkyl (e.g.        methyl), (C₆-C₁₈)aryl (e.g. phenyl), (C₁-C₁₂)acyl (formyl,        acetyl, napthaleneacetyl, and myristoyl), (C₇-C₁₈)aralkyl (e.g.        benzyl), and (C₇-C₁₈)alkaryl (e.g. p-methlyphenyl);-   A²² is an aromatic amino acid, Ala, Aib, Anb, N-Me-Ala or is    deleted;-   A²³ is Ser, Thr, Ala, Aib, N-Me-Ser, N-Me-Thr, N-Me-Ala or is    deleted;-   A²⁴ is Leu, Ile, Nle, Val, Trp, Gly, Nva, Aib, Anb, N-Me-Leu or is    deleted;-   A²⁵ is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-pε-NH—R (where R is    H, a branched or straight chain (C₁-C₁₀) alkyl group, or an aryl    group), Orn or is deleted;-   A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His, β-pyrazolylalaline,    N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-ε-NH—R (where R    is H, a branched or straight chain (C₁-C₁₀) alkyl group, an aryl    group, or a pharmaceutically acceptable salt thereof), Orn or is    deleted;-   A²⁷ is an aromatic amino acid;-   A²⁸ is Leu, Ile, Nle, Val, Trp, Aib, Anb or N-Me-Leu;-   A²⁹ is Asn, Ala, Gln, Gly, Trp or N-Me-Asn;-   A³⁰ is Leu, Ile, Nle, Nva, Fla, Val, Trp, Aib, Anb or N-Me-Leu;-   A³¹ is Val, Leu, Nle, Nva, Ile, Trp, Aib, Anb or N-Me-Val;-   A³² is Thr, Ser, D-Trp, N-Me-Ser or N-Me-Thr; and    wherein the compound optionally includes one or two pseudopeptide    bonds where each pseudopeptide bond is independently selected from    —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.

A preferred compound of formula (IV) includes N-α-Ac[Trp²⁷,ψ^(35/36)]PYY(25-36)-NH₂ (SEQ. ID. NO. 5), referred to as BT-56,N-α-Ac[Trp³⁰]PYY(25-36)-NH₂ (SEQ. ID. NO. 4), referred to as BWX-115,and N-α-Ac-[Trp³⁰, ψ^(35/36)]PYY(25-36)-NH₂ (SEQ. ID. NO. 6), referredto as BT-123, wherein ψ in the foregoing formulas is —CH2-NH—. Apreferred compound of formula (V) includes N-α-Ac[Nle^(24,28), Trp³⁰,Nva³¹, ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO. 7), referred to as B-48,wherein ψ is —CH2-NH—.

The formulas of I-V, as indicated above, optionally include at least onepseudopeptide bond between amino acid residues. By “psuedopeptide bond”is meant that the carbon atom participating in the bond between tworesidues is reduced from a carbonyl carbon to a methylene carbon, i.e.,CH₂—NH; or less preferably that of CO—NH is replaced with any of CH₂—S,CH₂—CH₂, CH₂—O, or CH₂—CO. A psuedopeptide bond may be symbolized hereinby “ψ”. The psuedopeptide bond analogs can be used to form dimericanalogs. A detailed discussion of psuedopeptide bonds is given in Coy etal. (1998) Tetrahedon 44:835-841.

In another aspect, the invention features a method of controlling thefood intake, i.e. appetite, of a subject comprising administering tosaid subject the compound of formula I, II, or III alone or incombination with formulas IV or V.

In other preferred embodiments, a therapeutically effective amount of acompound of formula I, II, or III alone, or in combination with formulasIV or V, and a pharmaceutically acceptable carrier substance togetherform a therapeutic composition capable of suppressing an NPY mediatedphysiological response.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a graph representing the feeding patterns of animals treatedwith a PP(32-36) analog by intraperitoneal injection. The compoundstested include a control (saline), hPP, and

FIG. 2 is a graph representing the feeding patterns of animals treatedwith a PP(32-36) analog (Y4 receptor agonist), and a mixture of thePP(32-36) analog and a PYY(22-36) analog (Y2 receptor agonist), byintraperitoneal injection. The compounds tested include a control(saline),

and a combination of B-74 and N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹,ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO. 7) (B-48) wherein ψ is —CH2-NH—;and

FIG. 3 is a graph representing the feeding patterns of animals treatedwith a PYY(22-36) analog (Y2 agonist receptor), and a mixture of thePYY(22-36) analog and a PP(32-36) analog (Y4 receptor agonist), byintraperitoneal injection. The compounds tested include a control(saline), N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹, ψ^(35/36)]PYY(22-36)-NH₂(SEQ. ID. NO. 7) (B-48) wherein ψ is —CH2-NH—, and a combination of B-48and

DETAILED DESCRIPTION OF THE INVENTION

The present invention is directed to neuropeptide Y (“NPY”) Y₄ receptoragonists including pancreatic polypeptide (PP), analogs thereof, andpeptide fragments of PP, e.g. PP(32-36), and analogs thereof, topharmaceutical compositions containing such Y₄ receptor agonists, and tomethods for treatment of mammals using the same. The NPY Y₄ receptoragonists may be administered to mammals either alone or in combinationwith NPY Y₂ receptor agonists including peptide YY (PYY) (3-36), analogsthereof, and to peptide fragments of PYY(3-36), e.g. PYY(22-36) andPYY(25-36), and analogs thereof, such as to control food intake inmammals, blood pressure, cardiovascular response, libido, circadianrhythm, hyperlipidimia, chronic pancreatitis, and nonalcoholic fattyliver disease including nonalcoholic steatohepatitis.

One advantage of using the Y4 selective ligands of the present inventionis that they can suppress food intake on peripheral administration,whereas Y1 and Y5 selective ligands, as described above, have topenetrate the BBB to modulate food intake. Additionally, an advantage ofusing Y2 selective ligands is that they also can suppress the foodintake on peripheral administration. Therefore, the inventor hasdeveloped NPY Y₄ receptor agonists that can be used alone or mixed withNPY Y₂ receptor agonists, as are disclosed in U.S. Pat. Nos. 5,604,203,and 6,046,167 to Balasubramaniam, which are herein incorporated byreference, for controlling an NPY mediated physiological response in asubject, such as to control food intake in mammals by suppressingappetite.

Accordingly, in one aspect, the present invention features NPY Y₄receptor agonists of PP(32-36) and analogs thereof, such pentapeptideanalogs having the formula:

wherein:

-   each X, independently, is an aromatic amino acid;-   each Y, independently, is an amino acid having a guanidino group;-   each Z, independently, is an aliphatic amino acid;-   n=1, 2, 3 or 4; and-   wherein the compound optionally includes one or two pseudopeptide    bonds where each pseudopeptide bond is independently selected from    —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.

A preferred compound of formula (I) includes,

(which is referred to as B-74).

In another aspect, the present invention features NPY Y₄ receptoragonists or pentapeptide analogs of PP(32-36) having the formula:

wherein:

-   each X, independently, is an aromatic amino acid;-   each Y, independently, is an amino acid having a guanidino group;-   each Z, independently, is an aliphatic amino acid;-   A1 and A2, independently, are selected from Cys, Pen, Glu, Asp, Lys,    and Dpr; and-   wherein the compound optionally includes one or two pseudopeptide    bonds where each pseudopeptide bond is independently selected from    —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.

A preferred compound of formula (II) includes,

In another aspect, the present invention features NPY Y₄ receptoragonists or pentapeptide analogs of PP(32-36) having the formula:H—[X—Y-Z-Y—X]_(n)—NH₂  (III)

wherein:

-   each X, independently, is an aromatic amino acid;-   each Y, independently, is an amino acid having a guanidino group;-   each Z, independently, is an aliphatic amino acid;-   n=1, 2, 3 or 4; and-   wherein the compound optionally includes one or two pseudopeptide    bonds where each pseudopeptide bond is independently selected from    —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.

Preferred compounds of formula (III) includeH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ. ID. NO. 9) orH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ.ID. NO. 10).

For formulas I-III, aromatic amino acids can include, for example,phenylalanine, tryptophan, or tyrosine, which is a hydroxyl aromaticamino acid; aliphatic amino acids can include, for example, isoleucine,leucine, norvaline, glycine, alanine, or valine; and amino acids thathave a guanidino group may include, for example, arginine. In addition,the C-terminal of the compounds of formulas (I-III) are amidated.

In another aspect, NPY Y₄ receptor agonists, such as those of formulas(III), are mixed or combined with NPY Y₂ receptor agonists, such asanalogs of PYY(25-36), for controlling an NPY mediated physiologicalresponse in a subject, e.g. to suppress appetite, such analogs ofPYY(25-36) having the formula:

wherein:

-   the N-terminal amino acid is bonded to R¹ and R²;-   Y is a chain of 0-4 amino acids, inclusive, where the C-terminal    amino acid has a carboxylamide group; which is independently bonded    to R³ and R⁴, e.g.,-   R¹ and R² are each independently bonded to the amino group of the    N-terminal amino acid and selected from H, (C₁-C₁₂)alkyl (e.g.    methyl), (C₆-C₁₈)aryl (e.g. phenyl), (C₁-C₁₂)acyl (formyl, acetyl,    napthaleneacetyl, and myristoyl), (C₇-C₁₈)aralkyl (e.g. benzyl), and    (C₇-C₁₈)alkaryl (e.g. p-methlyphenyl);-   R³ and R⁴ are each independently bonded to the amide group of the    C-terminus amino acid, e.g.,    -   (where R denotes the side chain group of the amino acid, e.g.        R═H in Gly, etc.), and selected from H, (C₁-C₁₂)alkyl (e.g.        methyl), (C₆-C₁₈)aryl (e.g. phenyl), (C₁-C₁₂)acyl (formyl,        acetyl, napthaleneacetyl, and myristoyl), (C₇-C₁₈)aralkyl (e.g.        benzyl), and (C₇-C₁₈)alkaryl (e.g. p-methlyphenyl);-   A²⁵ is Arg, Lys, homo-Arg, diethyl-homo-Arg, lys-ε-NH—R (where R is    H, a branched or straight chain (C₁-C₁₀) alkyl group, or an aryl    group), Orn or is deleted;-   A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His, β-pyrozolylalanin,    N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-e-NH—R (where R    is H, a branched or straight chain (C₁-C₁₀) alkyl group, or an aryl    group), Orn or is deleted;-   A²⁷ is an aromatic amino acid:-   A²⁸ is Leu, Ile, Val, Trp Nle, Nva, Aib, Anb, or N-Me-Leu;-   A²⁹ is Asn, Ala, Gin, Fly, Trp, or N-Me-Asn;-   A³⁰ is Leu, Ile, Val, Trp, Nle, Nva, Aib, Anb, or N-Me-Leu;-   A³¹ is Val, Ile, Trp, Nva, Aib, Anb, or N-Me-Val;-   A³² is Thr, Ser, N-Me-Ser, N-Me-Thr, or D-Trp.

In preferred embodiments Y is A³³-A³⁴-A³⁵-A³⁶ wherein

A³³ is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-ε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) alkyl group, or (C₆-C₁₈) arylgroup), Cys, or Orn

A³⁴ is Gln, Asn, Ala, Gly, N-Me-Gin, Aib, Cys, or Anb;

A³⁵ is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-ε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) alkyl group, or (C₆-C₁₈) arylgroup), Cys, or Orn; and

A³⁶ is an aromatic amino acid, Cys, or a pharmaceutically acceptablesalt thereof.

Most preferably, the compound of formula (IV) includes N-α-Ac[Trp²⁷,ψ^(35/36)]PYY(25-36)-NH₂ (SEQ. ID . NO. 5), referred to as BT-56, andN-α-Ac[Trp³⁰]PYY(25-36)-NH₂ (SEQ. ID. NO. 4), referred to as BWX-115,and N-α-Ac-[Trp³⁰, ψ^(35/36)]PYY(25-36)-NH₂ (SEQ. ID. NO. 6), referredto as BT-123, wherein ψ in the foregoing formulas is —CH2-NH—.

In yet another aspect, NPY Y₄ receptor agonists, such as those offormulas (I-III), are mixed with NPY Y₂ receptor agonists, such asanalogs of PYY(22-36), for controlling an NPY mediated physiologicalresponse in a subject, e.g. to suppress appetite, such analogs ofPYY(22-36) having the formula:

wherein

-   X is a chain of 0-5 amino acids, inclusive, where the N-terminal    amino acid is bonded to R¹ and R² by the nitrogen of the amino group    of the N-terminal amino acid;-   Y is a chain of 0-4 amino acids, inclusive, where the C-terminal    amino acid has a carboxylamide group; which is independently bonded    to R³ and R⁴, e.g.,-   R¹ and R² are each independently bonded to the amino group of the    N-terminal amino acid and selected from H, (C₁-C₁₂)alkyl (e.g.    methyl), (C₆-C₁₈)aryl (e.g. phenyl), (C₁-C₁₂)acyl (formyl, acetyl,    napthaleneacetyl, and myristoyl), (C₇-C₁₈)aralkyl (e.g. benzyl), and    (C₇-C₁₈)alkaryl (e.g. p-methlyphenyl);-   R³ and R⁴ are each independently bonded to the amide group of the    C-terminus amino acid, e.g.,    -   (where R denotes the side chain group of the amino acid, e.g.        R═H in Gly, etc.), and selected from H, (C₁-C₁₂)alkyl (e.g.        methyl), (C₆-C₁₈)aryl (e.g. phenyl), (C₁-C₁₂)acyl (formyl,        acetyl, napthaleneacetyl, and myristoyl), (C₇-C₁₈)aralkyl (e.g.        benzyl), and (C₇-C₁₈)alkaryl (e.g. p-methlyphenyl);-   A²² is an aromatic amino acid, Ala, Aib, Anb, N-Me-Ala or is    deleted;-   A²³ is Ser, Thr, Ala, Aib, N-Me-Ser, N-Me-Thr, N-Me-Ala or is    deleted;-   A²⁴ is Leu, Ile, Nle, Val, Trp, Gly, Nva, Aib, Anb, N-Me-Leu or is    deleted;-   A²⁵ is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-pε-NH—R (where R is    H, a branched or straight chain (C₁-C₁₀) alkyl group, or an aryl    group), Orn or is deleted;-   A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His, β-pyrazolylalaline,    N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-ε-NH—R (where R    is H, a branched or straight chain (C₁-C₁₀) alkyl group, an aryl    group, or a pharmaceutically acceptable salt thereof), Orn or is    deleted;-   A²⁷ is an aromatic amino acid;-   A²⁸ is Leu, Ile, Nle, Val, Trp, Aib, Anb or N-Me-Leu;-   A²⁹ is Asn, Ala, Gln, Gly, Trp or N-Me-Asn;-   A³⁰ is Leu, Ile, Nle, Nva, Fla, Val, Trp, Aib, Anb or N-Me-Leu;-   A³¹ is Val, Leu, Nle, Nva, Ile, Trp, Aib, Anb or N-Me-Val; and-   A³² is Thr, Ser, D-Trp, N-Me-Ser or N-Me-Thr.

In preferred embodiments, Y is A³³-A³⁴-A³⁵-A³⁶ wherein

A³³ is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-ε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) alkyl group, or an aryl group), Cys,or Orn

A³⁴ is Cys, Gln, Asn, Ala, Gly, N-Me-Gin, Aib, or Anb;

A³⁵ is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-ε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) alkyl group, or an aryl group), Cys,or Orn; and

A³⁶ is an aromatic amino acid, Cys, or a pharmaceutically acceptablesalt thereof.

Most preferably, the compound of formula (V) includesN-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹, ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO.7), referred to as B-48, wherein ψ is —CH2-NH—.

Concerning formulas IV and V, reference to the (C₁-C₁₂)acyl for R1 andR2 can include an aliphatic acyl (e.g. CH₃CO) or an aromatic acyl (e.g.C₆H₅CO).

Additionally, the formulas of I-V optionally include at least onepseudopeptide bond between amino acids residues. By “psuedopeptide bond”is meant that the carbon atom participating in the bond between tworesidues is reduced from a carbonyl carbon to a methylene carbon, i.e.,CH2-NH; or less preferably that of CO—NH is replaced with any of CH₂—S,CH₂—CH₂, CH₂—O, or CH₂—CO. A psuedopeptide peptide bond is symbolizedherein by “ψ”. Preferably, the psuedopeptide bonds are located betweenone or more amino acid residues. In addition, such psuedopeptide bondanalogs can be used to form dimeric analogs. A detailed discussion ofpsuedopeptide bonds is given in Coy et al. (1998) Tetrahedon 44:835-841.

Accordingly, in another aspect and as already specifically illustratedby certain of the preferred compounds, the invention features dimers,trimers, etc. of compounds having the formula (I-V). These dimers,trimers, etc. may be prepared, for example, by dimerizing compounds offormula (I-V) with dicarboxylic acids (e.g., succinic acid), cystine, ordiaminodicarboxylic acid (e.g., 2,6-diaminopimelic acid) as is known inthe art. Resulting compounds can include, for example, pentapeptidelinear tandem or parallel dimers.

In another aspect, the invention features a method of controlling thefood intake, i.e. appetite, of a subject comprising administering tosaid subject the compound of formula I, II, or III alone or incombination with formulas IV or V. These compounds also can beconjugated to carriers, such carriers can include albumin, such ascationized albumin (Endocrinology 126:977-984 (1990); J. Pharmacol Exp.Therao. 268:791-796 (1994)), polyethylene glycol, polyglutamic acid,polyleucine, polyisoleucine or polylysine, e.g., MAP.

In other preferred embodiments, a therapeutically effective amount of acompound of formula I, II, or III alone, or in combination with formulasIV or V, and a pharmaceutically acceptable carrier substance, e.g.,magnesium carbonate or lactose, together form a therapeutic compositioncapable of suppressing an NPY mediated physiological response. Thiscomposition can be in the form of a pill, tablet, capsule, liquid, orsustained released tablet for oral administration; or a liquid for nasaladministration as drops or spray; or a liquid for intravenous,subcutaneous, parenteral, or intraperitoneal administration. Anotherpreferred form for administration includes a biodegradablesustained-release composition for intramuscular administration to asubject in need of the composition. Preferably, the composition includesa lipophilic salt and is suitable for administration in the form of anoil emulsion or dispersion to a subject in need of the composition.

In yet another aspect, the invention features methods for controlling anNPY mediated physiological response in a subject; such methods involveadministering a compound of formula I, II, or III alone, or incombination with formulas IV or V, to a subject in a dosage effective tocontrol blood pressure, cardiovascular response, libido, circadianrhythm, hyperlipidimia, chronic pancreatitis, and nonalcoholic fattyliver disease including nonalcoholic steatohepatitis.

The NPY Y₄ receptor agonists including pancreatic polypeptide (PP),analogs thereof, and peptide fragments of PP, e.g. PP(32-36), andanalogs thereof such as those of compounds (I-III), either alone or incombination with NPY Y₂ receptor agonists including peptide YY (PYY)(3-36), analogs thereof, and to peptide fragments of PYY(3-36), e.g.PYY(22-36) and PYY(25-36), and analogs thereof such as those ofcompounds (IV-V), also can be useful in treating any number of illnessesthat involve eating disorders, cardiovascular function, alterations insexual function, as well as disorders of sleep and circadian rhythms(see, e.g., Harrison's Principles of Internal Medicine, McGraw-HillInc., New York, 12th ed.).

Other features and advantages of the invention will be apparent to oneskilled in the art.

The symbol A1, A2, A3, and the like; and Tyr, Lys or the like, as foundin a peptide sequence herein stands for an amino acid residue, e.g.,—N—CH(R)—CO— when it is at the N-terminus, or —NH—CH(R)—CO— when it isat any other position, where R denotes the side chain (or identifyinggroup) of an amino acid or its residue. For example, R is CH₂COOH forAsp, R is —H for Gly, R is —CH₂OH for Ser, R is —CH₃ for Ala and R is—CH₂CH₂CH₂CH₂NH₂ for Lys.

As set forth above and for convenience in describing this invention, theconventional and certain unconventional abbreviations for the variousamino acids are used. They are familiar to those skilled in the art; butfor clarity are listed below. All peptide sequences mentioned herein arewritten according to the usual convention whereby the N terminal aminoacid is on the left and the C-terminal amino acid is on the right. Ashort line between two amino acid residues indicates a peptide bond.

Abbreviations (common):

Asp=D=Aspartic Acid

Ala=A=Alanine

Arg=R=Arginine

Asn =N=Asparagine

Cys =C=Cysteine

Gly=G=Glycine

Glu=E=Glutamic Acid

Gin=Q=Glutamine

His =H=Histidine

Ile=I=Isoleucine

Leu=L=Leucine

Lys=K=Lysine

Met=M=Methionine

Phe =F=Phenylalanine

Pro=P=Proline

Ser=S=Serine

Thr=T=Threonine

Trp=W=Tryptophan

Tyr=Y=Tyrosine

Val=V=Valine.

Abbreviations (uncommon):

Aoc=8-aminooctanoic acid

Orn=Ornithine

Nal=2-napthylalanine

Thi=2-thienylalanine

Pcp=4-chlorophenylalanine

Bth=3-benzothienyalanine

Bip=4,4′-biphenylalanine

Tic=tetrahydroisoquinoline-3-carboxylic acid

Aib=aminoisobutyric acid

Anb=alpha-aminonormalbutyric acid

Ac₆c=1-aminocyclohexanecarboxylic acid

D-Pal=beta-(3-pyridyl)alanine;

Tcc=tetrahydrocarbolenecarboxylic acid

Abu=α-aminonormalbutyric acid

hArg(Pr)₂=N,N′-guanidino-(dipropyl)-homoarginine

Tic-OH=1,2,3,4 tetrahydroisoquinoline-7-hydroxy-3-carboxylic acid

Dip=3,3-diphenylalanine

2-Nal=3(2-naphthylalanine)

Tfp=Paratrifluoromethyl phenylalanine

Fla=3-(9-Fluorenyl)alanine

Flg=9-Fluorenylglycine

Cit=Citruline

Adp=2,5-diaminoadipic acid

Pim=2,6-diaminopimelicacid

Sub=2,7-diaminosuberic acid

Nle=Norleucine

Nva=Norvaline

Thz=4-Thiazolylalanine

Dpr=Dap=2,3-diaminopropionic acid

Pyr=Pyroglutamic acid

Tip=1,2,3,4-tetrahydronorhamman-3-carboxylic acid

Pen=Penicillamine.

Other Abbreviations:

Fmoc =N-(9-fluorenyl)methoxycarbonyl.

The terms “C2-C4-alkenyl” and “C2-C6-alkenyl” as used herein refer to a2 to 4 to 6 straight- or branched-chain of carbon atoms which contains acarbon-carbon double bond, such as allyl, propenyl, butanol, isoprenyland the like.

The terms “C1-C18-alkyl” as used herein refer to straight or branchedchain alkyl radicals having from 1 to 18 carbon atoms including, but notlimited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl,sec-butyl, pentyl, neopentyl hexyl, and the like.

The term “C6-C18-aryl” as used herein refers to phenyl or to a “bicycliccarbocyclic” group or “bicyclic carbocycle” having two fused carbocyclicrings, each ring having 5, 6 or 7 carbon atoms, and each ring beingfully saturated, partially saturated or aromatic. Bicyclic carbocyclicgroups include, but are not limited to, naphthyl, tetrahydronaphthyl,decalin, indanyl, indenyl and the like.

The term “C7-C18-arylalkyl” as used herein refers to an aryl groupappended to a C1-C4-alkyl radical including, but not limited to, benzyl,phenethyl, naphthylmethyl and the like.

The term “bicyclic heterocycle” as used herein refers to a group havingtwo fused rings, one or both of which are heterocyclic rings as definedherein. When both rings are not heterocyclic, the other ring iscarbocyclic and is saturated, partially saturated or aromatic,preferably a benzene ring. Bicyclic heterocyclic groups can beunsubstituted or monosubstituted or disubstituted with substituentsindependently selected from hydroxy, halo, oxo (═O), amino,C1-C4-alkylamino, di-(C1-C4)-alkylamino, C1-C4-alkoxy,thio-C1-C4-alkoxy, carboxy, C1-C4-alkoxycarbonyl, C1-C4-alkyl,C3-C8-cycloalkyl, —OSO₃H and halo-C1-C4-alkyl. Examples of bicyclicheterocycles include indole, 5-hydroxyindole, quinoline, isoquinoline,tetrahydroisoquinoline, quinoxaline, benzimidazole, benzofuran, and thelike.

The term “cyclo-C3-C10-alkyl” as used herein refers to an aliphaticmonocyclic of 3 to 10 or bicyclic group having 6 to 10 carbon atomsincluding, but not limited to, cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl, cyclooctyl, adamantyl, and the like.

The term “halo” or “halogen” as used herein refers to chloro, bromo,iodo or fluoro.

The term “halo-C1-C4-alkyl” as used herein refers to a lower alkylradical in which one to three hydrogen atoms have been replaced by ahalogen including, but not limited to, chloromethyl, 2-fluoroethyl,trifluoromethyl and the like.

The term “monocyclic heterocyclic group” or “monocyclic heterocycle” asused herein refers to any 3- or 4-membered ring containing a heteroatomselected from oxygen, nitrogen and sulfur, or a 5- or 6-membered ringcontaining carbon atoms and one, two or three nitrogen atoms; onenitrogen and one sulfur atom; or one nitrogen and one oxygen atom;wherein the 5-membered ring has 0-2 double bonds and the 6-membered ringhas 0-3 double bonds; wherein the nitrogen and sulfur heteroatoms mayoptionally be oxidized; and wherein the nitrogen heteroatom mayoptionally be quaternized. Heterocycles include, but are not limited to,pyridyl, imidazolyl, furyl, thienyl, pyrazinyl, pyrrolyl, pyrimidyl andthe like. Heterocyclics may be unsubstituted or mono- or disubstitutedwith substituents independently selected from hydroxy, halo, oxo (═O),amino, C1-C4-alkylamino, (C1-C4)-2-alkylamino, C1-C4-alkoxy,thio-C1-C4-alkoxy, carboxy, C1-C4-alkoxycarbonyl, C1-C4-alkyl,C3-C8-cycloalkyl, —OSO₃H and halo-C1-C4-alkyl.

The term “peptide bond” as used herein refers to the chemical bondbetween carbon and nitrogen in the bivalent group CONH that unites aminoacid residues in a peptide.

In addition, the above compounds may contain two or more asymmetriccarbon atoms and thus can exist as pure diastereomers, mixtures ofdiastereomers, diastereomeric racemates or mixtures of diastereomericracemates. As such, the present invention includes within its scope allof the isomeric forms. In keeping with standard peptide nomenclature, J.Biol. Chem., 1969, 243:3557-59, abbreviations for amino acid residuesare used herein.

It is noted that all amino acid residue sequences are represented hereinby formulae whose left to right orientation is in the conventionaldirection of amino-terminus to carboxy-terminus.

Administration

The amount of active ingredient that may be combined with the carriermaterials to produce a single dosage form will vary depending upon thehost treated, the particular treatment and the particular mode ofadministration.

It will be understood, however, that the specific dose level for anyparticular patient will depend upon a variety of factors including theactivity of the specific compound employed, the age, body weight,general health, sex, diet, time of administration, rate of excretion,drug combination, and the severity of the particular disease undergoingtherapy.

However, generally speaking the following guidelines will suffice. Whenan NPY Y₄ receptor agonist, such as those of compounds (I-III), iseither used alone or in combination with an NPY Y₂ receptor agonist,such as those of compounds (IV-V), as an agonist(s) of NPY in a humansubject, the total daily dose administered in single or divided dosesmay be in amounts, for example, from 0.001 to 1000 mg a day and moreusually 1 to 1000 mg. Dosage unit compositions may contain such amountsof submultiples thereof to make up the daily dose.

The compounds useful in the present inventive method may be administeredby any suitable means. One skilled in the art will appreciate that manysuitable methods of administering the compounds to an animal in thecontext of the present invention, in particular a human, are available,and, although more than one route may be used to administer a particularcompounds, a particular route of administration may provide a moreimmediate and more effective reaction than another route.

The compositions according to the present invention may be formulatedfor administration by any suitable route such as the oral, rectal,nasal, topical (dermal) or parenteral administration route. Thus, thecomposition may be in the form of tablets, capsules, suspensions,emulsions, solutions, injectables, suppositories, sprays, aerosols andin other suitable form.

Formulations for oral use include tablets that contain the activeingredient in admixture with non-toxic pharmaceutically acceptableexcipients. These excipients may be, for example, inert diluents, suchas calcium carbonate, sodium chloride, lactose, calcium phosphate orsodium phosphate; granulating and disintegrating agents, for example,potato starch or alginic acid; binding agents, for example, starch,gelatin or acacia; and lubricating agents, for example, magnesiumstearate, stearic acid or talc. Other pharmaceutically acceptableexcipients can be colorants, flavoring agents, plasticizers, humectantsetc. The tablets may be uncoated or they may be coated by knowntechniques, optionally to delay disintegration and absorption in thegastrointestinal tract and thereby provide a sustained action over alonger period. For example, a time delay material such as glycerylmonostearate or glyceryl distearate may be employed.

Formulations for oral use may also be presented as chewing tablets, oras hard gelatin capsules wherein the active ingredient is mixed with aninert solid diluent, for example, calcium carbonate, calcium phosphateor kaolin, or as soft gelatin capsules wherein the active ingredient ismixed with water or an oil medium, for example, peanut oil, liquidparaffin, or olive oil.

Powders, dispersible powders or granules suitable for preparation of anaqueous suspension by addition of water are also convenient dosage formsof the present invention. Formulation as a suspension provide the activeingredient in admixture with a dispersing or wetting agent, suspendingagent and one or more preservatives. Suitable dispersing or wettingagents are, for example, naturally-occurring phosphatides, as e.g.lecithin, or condensation products of ethylene oxide with e.g. a fattyacid, a long chain aliphatic alcohol or a partial ester derived fromfatty acids and a hexitol or a hexitol anhydrides, for example,polyoxyethylene stearate, polyoxyethylene sorbitol monooleate,polyoxyethylene sorbitan monooleate etc. Suitable suspending agents are,for example, sodium carboxymethylcellulose, methylcellulose, sodiumalginate etc.

The present agents can also be administered in the form of liposomes. Asis known in the art, liposomes are generally derived from phospholipidsor other lipid substances. Liposomes are formed by mono- ormulti-lamellar hydrated liquid crystals that are dispersed in an aqueousmedium. Any non-toxic, physiologically acceptable and metabolizablelipid capable of forming liposomes can be used. The present compositionsin liposome form can contain, in addition to the peptides of the presentinvention, stabilizers, preservatives, excipients, and the like. Thepreferred lipids are the phospholipids and the phosphatidyl cholines(lecithins), both natural and synthetic.

Methods to form liposomes are known in the art. See, for example,Prescott, Ed., Methods in Cell Biology, Volume XIV, Academic Press, NewYork, N.Y. (1976), p. 33 et seq.

The pharmaceutical formulation may also be administered parenterally(intravenous, intramuscular, subcutaneous or the like) in dosage formsor formulations containing conventional, non-toxic pharmaceuticallyacceptable carriers and adjuvants. The formulation and preparation ofsuch compositions is well-known to those skilled in the art ofpharmaceutical formulation.

For parenteral use, the pharmaceutical compositions according to theinvention may comprise the thermogenic compounds in the form of asterile injection. To prepare such a composition, the compounds aredissolved or suspended in a parenterally acceptable liquid vehicle.Among acceptable vehicles and solvents that may be employed are water,water adjusted to a suitable pH by addition of an appropriate amount ofhydrochloric acid, sodium hydroxide or a suitable buffer,1,3-butanediol, Ringer's solution and isotonic sodium chloride solution.The aqueous formulation may also contain one or more preservatives, forexample, methyl, ethyl or n-propyl p-hydroxybenzoate.

For the rectal application, suitable dosage forms for a compositionaccording to the present invention include suppositories (emulsion orsuspension type), and rectal gelatin capsules (solutions orsuspensions). In a typical suppository formulation, the compounds arecombined with an appropriate pharmaceutically acceptable suppositorybase such as cocoa butter, esterified fatty acids, glycerinated gelatin,and various water-soluble or dispersible bases like polyethylene glycolsand polyoxyethylene sorbitan fatty acid esters. Various additives like,for example, enhancers or surfactants, may be incorporated.

For the nasal application, typical dosage forms for a compositionaccording to the present invention include nasal sprays and aerosols forinhalation. In a typically nasal formulation, the active ingredients aredissolved or dispersed in a suitable vehicle. The pharmaceuticallyacceptable vehicles and excipients and optionally other pharmaceuticallyacceptable materials present in the composition such as diluents,enhances, flavoring agents, preservatives, etc., are all selected inaccordance with conventional pharmaceutical practice in a mannerunderstood by the persons skilled in the art of formulatingpharmaceuticals.

The pharmaceutical compositions according to the invention may also beadministered topically on the skin for percutaneous absorption in dosageforms or formulations containing conventionally non-toxicpharmaceutically acceptable carriers and excipients includingmicrospheres and liposomes. The formulations include creams, ointments,lotions, liniments, gels, hydrogels, solutions, suspensions, pastes,plasters and other kinds of transdermal drug delivery systems. Thepharmaceutically acceptable carriers or excipients may includeemulsifying agents, antioxidants, buffering agents, preservatives,humectants, penetration enhancers, chelating agents, gel-forming agents,ointment bases, perfumes and skin protective agents.

Examples of emulsifying agents are naturally occurring gums, e.g., gumacacia or gum tragacanth, naturally occurring phosphatides, e.g.,soybean lecithin and sorbitan monooleate derivatives.

Examples of antioxidants are butylated hydroxy anisole (BHA), ascorbicacid and derivatives thereof, tocopherol and derivatives thereof andcysteine.

Examples of preservatives are parabens and benzalkonium chloride.

Examples of humectants are glycerin, propylene glycol, sorbitol andurea.

Examples of penetration enhancers are propylene glycol, DMSO,triethanoiamine, N,N-dimethylacetamide, N,N-dimethylformamide,2-pyrrolidone and derivatives thereof, tetrahydrofurfuryl alcohol andAZONE®.

Examples of chelating agents are sodium EDTA, citric acid and phosporicacid.

Examples of gel forming agents are Carbopol, cellulose derivatives,bentonit, alginates, gelatin and PVP.

Examples of ointment bases are beeswax, paraffin, cetyl palmitate,vegetable oil, sorbitan esters of fatty acids (Span),polyethyleneglycols, and condensation products between sorbitan estersof fatty acids and ethylene oxide, e.g., polyoxyethylene sorbitanmonooleate (Tween).

The formulation and preparation of the above-mentioned compositions iswell-known to those skilled in the art of pharmaceutical formulation.Specific formulation can be found in “Remington's PharmaceuticalSciences” incorporated herein by reference.

In one aspect the present invention relates to a method for treatment ofoverweight or obesity in individuals, in particular in humans or forreducing the adipose tissue mass/lean mass body mass ratio of anindividual, in particular a human or a domestic animal.

In the present context the term “overweight” is used as an indication ofa body with a weight exceeding the “desirable weight”, whereas the term“obesity” is used when the body weight is 20% or more above the“desirable weight”. Desirable weights for humans are given by theCouncil on Scientific Affairs defining the desirable weights for humansaccording to Metropolitan Height and Weight Tables as the midpoint ofthe range of the medium-frame individuals.

In another aspect, the present invention relates to a method for thetreatment of diseases that are complications to overweight or obesity.These diseases or conditions include, for example, diabetes mellitustype II, hypercholesterolemia, hypertriglyceridaemia and hypertension.

In another aspect, the present invention also relates to a method ofreducing adipose tissue mass/lean body mass ratio or treating overweightor obesity or complications thereof by means of subjecting theindividuals to a diet regimen. The diet regimen into which theindividuals may be subjected in connection with the administration ofcompositions of the present invention may include a low carbohydrate, alow fat and a low energy regimen, e.g., a diet of from 800-2500kcal/day.

Veterinary Use

The NPY Y₄ receptor agonists including pancreatic polypeptide (PP),analogs thereof, and peptide fragments of PP, e.g. PP(32-36), andanalogs thereof such as those of compounds (I-III), either alone or incombination with NPY Y₂ receptor agonists including peptide YY (PYY)(3-36), analogs thereof, and to peptide fragments of PYY(3-36), e.g.PYY(22-36) and PYY(25-36), and analogs thereof such as those ofcompounds (IV-V), can also be administered to domestic animals in orderto improve the performance of the animal (daily weight gain and feedutilization) or to improve carcass quality or both. Carcass quality isgenerally improved when the fat tissue mass/lean mass body mass ratio isdecreased, i.e., when the body content of meat is increased e.g., at theexpense of the body content of fat.

The improvements in performance and carcass quality are suggested to becaused by a reduced fat accretion and/or by an increased skeletal muscleaccretion. In growing animals, the amount of lipid present is suggestedto be governed by the relative rates of lipolysis and lipogenesis.Stimulation of lipolysis and/or inhibition of lipogenesis in fat tissuemay lead to a reduced fat accretion. In vivo and in vitro studies withboth pigs and ruminants may indicate that certain beta agonistsstimulate lipolysis and inhibit lipogenesis in fat tissue leading to areduced fat accretion.

Administration to an animal of the compositions according to the presentinvention may be useful in order to increase the lean body mass at theexpense of body fat, particularly in domestic animals like pigs, hogs,cattle, sheep and poultry. The composition may be given in admixturewith the feed in a suitable dose corresponding to the size of theanimal.

Peptide Synthesis

The peptides of the present invention can be synthesized by any suitablemethod, such as by exclusively solid-phase techniques, by partialsolid-phase techniques, by fragment condensation or by classicalsolution addition. A detailed description of certain of these methods iscontained in “The Peptides, Vol. 1”, Gross and Meinenhofer, Eds.,Academic Press, New York, 1979. Coupling methods employed include thecarbodiimide method (1,3-dicyclohexylcarbodiimide [DCC],1-(3-dimethylaminopropyl-3-ethylcarbodiimide hydrochloride [EDCI]) withthe option of racemization preventing additives (1-hydroxybenzotriazole[HOBT]), the mixed anhydride method, the azide method, the acid chloridemethod, the symmetrical anhydride method, the use ofbis(2-oxo-3-oxazolidinyl)phosphinic chloride (BOP—Cl), and the activeester method (N-hydroxysuccinimide esters, 4-nitrophenol esters,2,4,5-trichlorophenol esters, and the like). The synthesis of the PYYanalogs of U.S. Pat. Nos. 5,604,203, and 6,046,167 to Balasubramaniam isfully disclosed therein, which is hereby incorporated by referenceherein, and further is generally discussed below.

The techniques of exclusively solid-phase synthesis are set forth in thetextbook “Solid-Phase Peptide Synthesis” Stewart & Young, Freeman & Co.,San Francisco, 1969, and are exemplified by the disclosure of U.S. Pat.No. 4,105,603, issued Aug. 8, 1978 to Vale et al. The fragmentcondensation method of synthesis is exemplified in U.S. Pat. No.3,972,859 (Aug. 3, 1976). Other available syntheses are exemplified byU.S. Pat. No. 3,842,067 (Oct. 15, 1974) and U.S. Pat. No. 3,862,925(Jan. 28, 1975). All of the above references are incorporated herein byreference.

Synthesis by the use of recombinant DNA techniques, for purposes of thisapplication, should be understood to include the suitable employment ofa structural gene coding for peptides, or peptide fragments, totransform a microorganism, using an expression vector including apromoter and operator together with such structural gene, and causingsuch transformed microorganism to express the peptide or such asynthetic peptide fragment. A non-human animal may also be used toproduce the peptide by gene-farming using such a structural gene in themicroinjection of embryos as described in U.S. Pat. No. 4,870,009 issuedSep. 26, 1989, incorporated herein by reference.

When the peptides are not prepared using recombinant DNA technology,they are preferably prepared using solid phase synthesis, such as thatdescribed by Merrifield, J. Am. Chem. Soc., 85, p 2149 (1964), althoughother equivalent chemical syntheses known in the art can also be used aspreviously mentioned. Solid-phase synthesis is commenced from theC-terminus of the peptide by coupling a protected alpha-amino acid to asuitable resin as generally set forth in U.S. Pat. No. 4,244,946 issuedJan. 21, 1981 to Rivier et al., the disclosure of which is incorporatedherein by reference. A starting material for analogs of the presentinvention can, for example, be prepared by attaching alpha-amino- andside-chain-protected Tyr to a BHA resin.

The compounds of the invention may be prepared by stepwise coupling ofthe amino acids or by coupling together fragments of dipeptide length orgreater. Thus, the free carboxylic acid moiety from one amino acid orpeptide fragment is activated and allowed to condense with the freenitrogen group of the second amino acid or peptide fragment. Thecoupling reactions are conducted in solvents such as methylene chloride(CH₂Cl₂), tetrahydrofuran (THF), dimethylformamide (DMF) or other suchsolvents.

During the coupling process, the non-participating carboxylic acids oramines on the reacting set of amino acids or peptide fragments areprotected by a protecting group that can be selectively removed at alater time if desired. A detailed description of these groups and theirselection and chemistry is contained in “The Peptides, Vol. 3”, Grossand Meinenhofer, Eds., Academic Press, New York, 1981, incorporatedherein in its entirety by reference. Thus, useful protective groups forthe amino group are benzyloxycarbonyl(Cbz), t-butyloxycarbonyl(t-BOC),2,2,2-trichloroethoxycarbonyl(Troc), t-amyloxycarbonyl,4-methoxybenzyloxycarbonyl, 2-(trichlorosilyl)ethoxycarbonyl,9-fluorenylmethoxycarbonyl(Fmoc), phthaloyl, acetyl (Ac), formyl,trifluoroacetyl, and the like.

Examples of useful protective groups for the carboxylic acid includeesters, such as methyl, ethyl, benzyl, t-butyl, 2,2,2-trichloroethyl,allyl, 4-nitrobenzyl, and the like. Removal of these protecting groupsmay be accomplished selectively by employing various acid or basecatalyzed hydrolytic, hydrogenolytic, thermal or dissolving metalconditions.

Generally, peptides will be synthesized by stepwise solid phasemethodology developed by using an automated Applied Biosystem Model 430Apeptide synthesizer. Tertiary butyloxy-carbonyl (Boc) amino acids withbenzyl or halobenzyl based side chain protecting groups (Asp & Glu withObzl or OcHex; Ser & Thr with Bzl: Cys with pMeBzl; Tyr with 2BrZ; Lyswith 2ClZ; Arg with Tos; His with Bom; Trp with CHO) will be used inconjunction with phenylacetamidomethyl (PAM) resin. In the case of thesynthesis of peptide amides, benzyldrylamine (BHA) orparamethylbenzylhydrylamine (MBHA) will be used instead of PAM resin.

Boc-aminoacid-PAM-resin, using Boc-aminoacyloxy-methyl-phenylacetic acidand aminomethyl resin, is available commercially. TheBoc-aminoacid-PAM-resin thus prepared eliminates the possibility ofchain termination by tri-fluoroacetylation. Attachment to BHA or MBHAresin will be performed by way of preformed symmetrical anhydride.

Coupling and deprotection functions are generally carried outautomatically by the instrument. The standard program provided by themanufacturers are modified to incorporate a double coupling procedure,first in DMF and then in CH₂Cl₂. Altering the polarity of the solventsimproves the coupling. All amino acids, except Asn, Gln and Arg, willgenerally be coupled as preformed symmetrical anhydrides. Asn, Gln andArg are double coupled as preformed 1-hydroxy-benzotriazole esters toavoid side reactions. Resin samples taken during these reactions may beassayed by quantitative procedure to determine the degree of coupling.Other standard cleaving reagents and conditions for removal of specificalpha-amino protecting groups may be used as described in Schroder &Lubke, “The Peptides” Vol. 1, pp 72-75 (Academic Press 1965),incorporated herein in its entirety by reference.

In the case of coupling unusual amino acids, suitable conditions(solubility, coupling times) will be first developed before using inautomated mode. In some cases these couplings will be carried outmanually (e.g.: pseudopeptides, N-Me-amino acids). Pseudopeptide bondswill be incorporated by the methods described earlier. The t-Boc-aminoacid aldehyde will be obtained by reducing N-methoxy-N-methylamidederivatives of Boc-amino acids with LiAlH₄. The aldehyde obtained willbe reacted immediately with the a-amino group of the peptide attached tothe resin in DMF containing 1.0% HOAC in the presence of an equivalentamount of NaBH₃CN. At the end of the reaction, the presence of secondaryamine is tested for with ninhydrin (wine-red color). The secondary amineformed will then be blocked by reacting with 2 equivalents ofZ(2-Cl)OSU, 2 equivalents of HOBT, and 4 equivalents ofdiisopropylethyamine until ninhydrin gives a yellow color. This way theformation of branched peptide is prevented. Coupling of stericallyhindered amino acids (e.g.: N-Me-amino acids, CαMeLeu, Aib) will beeffected by a HOAT or HATU which has been shown to be superior toBOP/HOBT.

For the final cleavage, the N-α-Boc group and the Nin-CHO will be firstremoved with 50% TFA/CH₂Cl₂ and 20% piperidine-DMF from the protectedpeptide resin before detaching the target peptide using HF containing p.cresol (5%). If Cys and Met are present, p. thicresol (2.5%) will alsobe added to the HF reaction mixture. If problems are encountered duringthe standard HF method, then the “low/high” HF procedure will be used.

The materials are then purified. After initial fractionation on SephadexG-25, the peptide material will be subjected to reversed phase highperformance liquid chromatography (RPLC) on C₁₈ Vydac columns. However,peptides may be first subjected to ion exchange chromatography beforeRPLC, depending upon the heterogeneity of the crude peptide. Thehomogeneity of the purified product may be confirmed by analytical RPLCusing two different solvent systems, amino acid analysis, completesequencing, and mass spectral analysis.

For analysis, the peptide resins are hydrolyxed using 12NHCl/HOAc/phenol (2:1:1) for 24 hours at 110° C. The free peptides arehydrolyzed for 24 hours in 6N HCl containing 0.1% phenol or 4N methanesulfonic acid at 110° C. and are quantified on a Waters Pico Tag system.Peptide hormones and fragments are then subjected to complete sequencingon an automated gas phase sequencer (Applied Biosystem, Model 470A).

For the production of a compound of the invention where any one orseveral of the constituent amino acids bear an N-alkyl group,specifically methyl, the corresponding N-alkyl amino acid can beprepared via the method described by Benoiton (Can. J. Chem., 1977,55:906) or Shuman (“Peptides: Proceedings of the 7th American PeptideSymposium”, D. Rich, E. Gross, Eds., Pierce Chemical Co., Rockford, Ill.1981, p 617), wherein the t-BOC- or Cbz-protected amino acid is treatedwith a base in the presence of a chelating agent such as a crown etherand then quenched with methyl iodide. An alternative method described byFreidinger (J. Org. Chem., 1983, 48:77), in which triethylsilanereduction of the oxazolidinone of an amino acid directly produces theN-methyl derivative may also be utilized.

The reduced carbonyl amide bond surrogates can be prepared in a mannersimilar to that described by Martinez (J. Med. Chem. 1987, 30:1366). TheN-alpha-t-BOC protected amino acid (with appropriate protection of sidechain functional groups) is converted to the 3,5-dimethylpyrazolide,which is then reduced with lithium aluminum hydride. The resultingaldehyde is then allowed to condense with an amino acid or peptidebearing a free amino terminus. Reduction of the Schiff base that isformed as a result of the condensation is accomplished using sodiumcyanoborohydride to yield the desired compound having a reduced amidebond.

Functionalization of the epsilon-amino group of the lysine (Lys) orhomologous (e.g., Orn) residue is achieved via activation of the acidfragment as the active ester (N-hydroxysuccinimide,2,4,5-trichlorophenol, etc.) or, if no other free carboxylic acidfunction is present on the peptide, coupling using any of the methodsmentioned above is applicable. In addition, the functionalization of theepsilon-amino group may be accomplished by reaction with various alkyland aryl isocyanates, as well as alkyl and aryl isothiocyanates.

The sulfuric acid esterification of the phenolic residues may beconducted using a variety of known reagents such as thepyridine-sulfuric anhydride or the pyridine-sulfur trioxide complex. Useof pyridinium acetyl sulfate as described by Penke and Rivier(“Proceedings of the 8th American Peptide Symposium”, V. Hruby, D. Rich,Eds., Pierce Chemical Company, Rockford, Ill.; 1983; p. 119), may alsobe applied to prepare the sulfuric acid ester derivative of thepeptides.

The compounds of the present invention can be used in the form of saltsderived from inorganic or organic acids. These salts include but are notlimited to the following: acetate, adipate, alginate, citrate,aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate,camphorsulfonate, digluconate, cyclopentanepropionate, dodecylsulfate,ethanesulfonate, glucoheptonate, glycerophosphate, hemisulfate,heptonate, hexanoate, fumarate, hydrochloride, hydrobromide,hydroiodide, 2-hydroxy-ethanesulfonate, lactate, maleate,methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate,pectinate, persulfate, 3-phenylpropionate, picrate, pivalate,propionate, succinate, tartrate, thiocyanate, tosylate, and undecanoate.Also, the basic nitrogen-containing groups can be quaternized with suchagents as loweralkyl halides, such as methyl, ethyl, propyl, and butylchloride, bromides, and iodides; dialkyl sulfates like dimethyl,diethyl, dibutyl, and diamyl sulfates, long chain halides such as decyl,lauryl, myristyl and stearyl chlorides, bromides and iodides, aralkylhalides like benzyl and phenethyl bromides, and others. Water oroil-soluble or dispersible products are thereby obtained.

The pharmaceutically acceptable salts of the present invention can besynthesized which contain a basic or acidic moiety by conventionalchemical methods. Generally, the salts are prepared by reacting the freebase or acid with stoichiometric amounts or with an excess of thedesired salt forming inorganic or organic acid or base in a suitablesolvent or various combinations of solvents.

Examples of preferred salts are those with therapeutically acceptableorganic acids, e.g., acetic, lactic, maleic, citirc, malic, ascorbic,succinic, benzoic, or pamoic acid, as wells as polymeric acids and saltswith inorganic acids such as the hydrohalic acids, e.g., hydrocholoricand sulfuric acids.

In addition, pseudopeptide bonds may, if desired, may be introduced atvarious positions, e.g., between amino acid residues A1-A2 or betweenresidues A2-A3. Optically pure Boc-AA-CHO can be obtained in good yieldsand coupled directly to the —NH2 group of the peptide resin by publishedmethods (Sasaki et al., Peptides 8:119-121, 1987; Fehrentz et al.,Synthesis pp. 676-678, 1983). The secondary amine in the pseudopeptidebond is capped with Z(2-Cl). This is introduced by reacting the peptideresin with Z(2-Cl)—OSU (2 equiv.), HOBT (2 equiv.) and DIEA (4 equiv.)for 10-60 min. The red wine color of ninhydrin with secondary amineturns yellow at the end of capping.

Exemplary compounds of the present invention include:

These compounds may be combined with NPY Y₂ receptor agonists includingPYY(3-36), analogs thereof, and peptide fragments of PYY(3-36), e.g.PYY(22-36) and PYY(25-36), and analogs thereof such as those of formulas(IV-V).

Other analogs of the invention can be prepared as above and tested fortheir biological activity effectiveness as antagonists or agonists usingthe methods described below and those commonly known in the art.

Functional Assays

Animals. Cell Lines and Cultures, And Reagents

Any suitable in vivo or in vitro system may be utilized to assay andtest the effectiveness of the compounds of the invention, andcombinations thereof as discussed above. Such assays may employ in vivomethods for evaluating physiological responses, e.g., blood pressure,renovascular function, feeding behavior, or circadian rhythm, or in vivobiochemical systems evaluating receptor binding in a suitable cell line,e.g., SK-N-MC (ATCC HBT 10) or SK-N-BE(2) (Barnes et al. In Vitro, 17:619 631, 1981); or in isolated cells, e.g., cells isolated from thespleen, kidney, heart or brain. A number of in vivo and in vitrobiochemical systems known to those skilled in the art are available fortesting antagonists to hypothalamic NPY receptors, e.g. the Y-1, Y-2,and Y-3 receptor categories. Described below are assay methods which canbe utilized with cell lines such as SK-N-MC and SK-N-BE2 or isolatedcardiac membranes which possess the high-affinity hypothalamic NPYreceptor sites. Other systems are also known for evaluating NPYantagonists to the hypothalamic NPY receptor, e.g. VSM cells (Sheikh etal., Am. J. Physiol. 260: G250 G257, 1991) and HEL cells (Motulsky etal. Amer. J. Physiol. 255: E880-E885, 1988); Y-2 receptor, e.g., kidney(Sheikh et al., Am. J. Physiol 26:F978-F984), spleen (Lunberg et al.,Eur. J. Pharmal. 145:21-29, 1988), dorsal root ganglion (Bleakman etal., Br. J. Pharmal. 103:1781-1789, 1991) and hippocampal cells (Sheikhet al., J. Biol. Chem. 265:8304 8310, 1990); and Y-3 receptors, e.g., incardiac ventricular membranes (Balasubramaniam et al., Peptides 11:545-550, 1990), chromaffin cells, rat gastric mucosa (Michel, M. C.,Trends in Pharmol. Sci. 12: 389-394, 1991) and brain stem.

In Vitro Biochemical Assays

The ability of the compounds of the invention, and combinations thereofas discussed above, to act as antagonists of NPY can be demonstrated byany number of methods known in the art. For example, the compounds canbe shown to compete with iodinated neuropeptide Y for receptors usingthe methods described by Lundberg et al. (Eur. J. Pharmol. 145: 21-29,1988); Gordon et al. (J. Neurochemistry 55:506-513, 1990); Walker et al.(Mol. Pharmacol. 34:779 792, 1988); Balasubramaniam et al. (Peptides10:1283-1286, 1989).

In one example demonstrating antagonists to hypothalamic NPY receptors,rat hypothalamus was isolated and the membranes were prepared forbinding and adenylate cyclase studies according to standard methods(Unden et al. 1984. Eur. J. Biochem 145: 525-530; Westlind-Danielsson etal., Neurosci. Lett. 74: 237-242 (1987)). Displacement studies areperformed in a total volume of 0.25 ml 20 mM HEPES buffer, pH 7.4,containing 1% bovine serum albumin, 0.1% bacitracin, 300 μm PMSF and 5KIU/ml aprotinin. In a standard assay, 100 μg of membrane/tube isincubated in a shaking water bath at 24° C. for 45 min with[¹²⁵I-Tyr]-NPY (20,000 CPM) as described by Balasubramaniam et al,(Peptides 11: 545-550, 1990), in the presence of increasingconcentrations of NPY (10 μOsM). At the end of incubation, 1.0 ml oficed cold buffer is added, centrifuged at 10,000×g for 10 min, and thesupernatant removed by aspiration. The tube containing the pellet iscounted for bound radioactivity in a micromedic gamma counter.

An example of assaying adenylate cyclase activity of hypothalamic andcerebral cortex membranes is now described.

Adenlyate cyclase activity of the hypothalamic and cerebral cortexmembranes is determined by incubating 50 μg of membranes in a totalvolume of 0.20 ml Tris-HCμ30 mM pH 7.4 buffer containing 150 mM NaCl,8.25 mM MgCl2, 0.75 mM EGTA, 1.5 theophylline, 20 μg/ml aprotinin, 100μg/ml bacitracin, 1 mg/ml bovine serume albumin, 1 mM ATP, 20 mMcreatine phosphate, 1 mg/ml phosphocreatine kinase, 10 μM isopreternol,10 μM GTP, and various concentrations of peptides (0-10 μM). Afterincubating the mixture at 35° C. for 15 min in a shaking water bath, thereaction is arrested by the addition of 100 μM EDTA and boiling for 3min. cAMP is extracted and quantitated by radioimmunoassay. All thepoints in the binding and adenlyate cyclase are the means of at leastthree parallel experiments performed in duplicate.

In Vivo Assays

Any suitable in vivo model system can be used to evaluate the propertiesof the compounds of the invention, and combinations thereof as discussedabove. Such models, without limitation, include those used to evaluatefeeding and memory behavior (Flood et al., Peptides 10:963-966), andvasoconstriction and hypertension (Balasubramaniam et al. Biochemm etBiophys Acta 997: 176-188, (1989)).

Thus, in one example, feeding studies can be performed using SpraqueDawley rats (350-450 g) with paraventricular hypothalmic cannulae toinvestigate effects of PP, NPY, and/or PYY analogs (Chance et al.,Peptides 10: 1283, 1286 (1989)).

The following Examples set forth preferred methods for synthesizing theanalogs of PP and of fragments thereof, such as the pentapeptide analogsof formulas (I-III) of the present invention, as well as the analogs ofPYY and of fragments thereof, such as those of formulas (IV-V), usingthe solid-phase technique which generally is in accordance with theprocedure set forth in U.S. Pat. No. 4,415,558 to Vale, et al., issuedNov. 15, 1983, the disclosure of which is incorporated herein byreference.

EXAMPLES Example I

Analogs of PP(32-36) and, more specifically, the pentapeptide paralleldimer,

which includes the tetra-methylene spacer, was synthesized generally bystandard stepwise t-Boc-solid phase methods and purified by reversedphase chromatography as is known in the art (J. Med. Chem. 2000, 49,3420-3427).

More specifically, to obtain the parallel dimer, B-74,N-α-Boc-D/L-diamino-dicarboxylic acids (0.5 equiv) was coupled manuallyto the N-terminus of the protected pentapeptide peptide resin in thepresence of equivalent quantities of DIC, HOBT, and DIEA. Free-peptideamides then were obtained by standard HF cleavage of the protectedpeptide-MBHA resin in the presence of about 5% p-cresol and about 2.5%thiocresol and purified by reversed phase chromatography.

Example II

To study the feeding patterns of animals treated with B-74, seven toeight week old C57BL/6 male mice (Harlan Laboratories, Indianapolis,Ind.) were individually housed in cages in a temperature controlled room(25° C.) under 12-hour light/dark cycles. Mice had free access to waterand standard chow and were acclimatized to daily ip saline injections.After one week of acclimatizing, mice were fasted for 18 hours beforethe experiment, and saline (0.1 ml), hPP (10 mmol/mice), or B-74 (100nmol/mice) in saline (0.1 ml) were injected intraperitoneally. Food wasprovided ten (10) minutes later, and the 2 and 4 hour food intake wasmonitored. Statistical significance was determined by ANOVA, withindividual means being compared post-hoc by Tukey's corrected t-test.

The results are shown in FIG. 1. Specifically, intraperitoneal (ip)injections of PP (10 nmol/mice) and B-74 (100 nmol/mice) inhibited the 2h and 4 h food intake to a greater extent than the control.

Example III

The PYY (22-36) analog, N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹,ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO. 7), referred to as B-48, whereinψ is —CH2-NH—, was similarly synthesized by standard stepwiset-Boc-solid phase methods and purified by reversed phase chromatographyas is known in the art according, and as referred to in Example I.

To further study the feeding patterns of animals treated with B-48,experiments were performed in seven to eight week old C57BL/6 male mice(Harlan Laboratories, Indianapolis, Ind.) after acclimatization of micefor one week. More specifically, in preliminary studies, overnight (18h) fasted animals were injected (ip) with 0.1 ml of saline or saline(0.1 ml) containing various concentrations of B-48 (0.025, 0.050, 0.100or 0.150 μmol/mice) (n=8-10 per group). The mice were provided with aknown quantity of chow 10 min later. The food intake was monitored over1-24 h. The results (not shown) revealed that a minimum dose of 0.050μmol/mice of B-48 was required to exhibit significant inhibitory effectson food intake.

Example IV

To further compare the inhibitory effects of injecting (ip) 0.050 μmolof B-48 or B-74 alone or together (B-48+B-74) on the food intake byfasted mice, additional experiments were conducted.

Accordingly, to study the feeding patterns of animals treated with B-48or B-74 alone or together (B-48+B-74), seven to eight week old C57BL/6male mice (Harlan Laboratories, Indianapolis, Ind.) were individuallyhoused in cages in a temperature controlled room (25° C.) under 12-hourlight/dark cycles. The mice had free access to water and standard chow.After acclimatizing for one week, mice were fasted 18 hours before theexperiment. The fasted animals were injected (ip) with 0.1 ml of salineor saline (0.1 ml) containing B-48 (0.050 μmol/mice) or B-74 (0.050μmol/mice) alone or together, B-48+B-74 (0.100 μmol/mice, i.e. 0.050μmol/mice per each compound), (n=8-10 per group). The mice were providedwith a known quantity of chow 10 minutes later. The food intake wasmonitored over 1-24 h. Statistical significance was determined by ANOVA,with individual means being compared post-hoc by Tukey's correctedt-test.

The results are shown in FIGS. 2 and 3. Specifically, treatment withB-74 or B-48 alone inhibited food intake to a greater extent than thecontrol over the 24 h time period, while the combination treatment(B-48+B-74) inhibited food intake to a greater extent than either B-48or B-74 taken alone. Table I below utilizes the data from FIGS. 2 and 3to further illustrate the percentage inhibition of food intake by B-74(0.05 μmol), B-48 (0.050 μmol), and B-74 (0.05 μmol)+B-48 (0.050 μmol)in the fasted mice at the 2 h, 4 h, 8 h, and 24 h marks. TABLE IPercentage of Inhibition of Food Intake Compared to Saline Hours afterInjection B-74 vs. (B-48 + B-74) B-48 vs. (B-48 + B-74) 2 35 vs. 85 39vs. 72 4 26 vs. 63 30 vs. 54 8 14 vs. 44 16 vs. 30 24  7 vs. 24  7 vs.20

Accordingly, as shown in FIGS. 2 and 3, and Table I, combined treatmentof B-48+B-74 caused greater inhibition of food intake at all time pointsas compared to individual treatment with either B-74 or B-48, or thecontrol.

While the present invention has been illustrated by the description ofthe various embodiments thereof, and while the embodiments have beendescribed in considerable detail, it is not intended to restrict or inany way limit the scope of the appended claims to such detail.Additional advantages and modifications will readily appeared to thoseskilled in the art. The invention in its broader aspects is thereforenot limited to the specific details, representative apparatus andmethods and illustrative examples shown and described. Accordingly,departures may be made from such details without departing from thescope of Applicants' general inventive concept. Various features of theinvention are emphasized in the claims that follow.

1. A compound having the formula:

wherein: each X, independently, is an aromatic amino acid; each Y,independently, is an amino acid having a guanidino group; each Z,independently, is an aliphatic amino acid; n=1, 2, 3 or 4; and whereinthe compound optionally includes one or two pseudopeptide bonds whereeach pseudopeptide bond is independently selected from —CH₂—NH—,—CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 2. The compound of claim 1wherein the compound includes one or two pseudopeptide bonds where eachpseudopeptide bond is —CH₂—NH—.
 3. The compound of claim 1 wherein thearomatic amino acid is selected from the group consisting of Phe, Trp,and Tyr and the aliphatic amino acid is selected from the groupconsisting of Ile, Leu, Val, Ala, Gly, and Nva.
 4. The compound of claim1 wherein the aromatic amino acid is a hydroxyl aromatic amino acid. 5.The compound of claim 1, wherein said compound is


6. A pharmaceutically acceptable salt of the compound of claim
 1. 7. Amethod for controlling an NPY mediated physiological response in asubject comprising administering to said subject the compound ofclaim
 1. 8. The method of claim 7 wherein the composition is capable ofsuppressing appetite.
 9. The method of claim 7 wherein the step ofadministering to said subject the compound of claim 1 includesadministering to said subject a mixture of the compound of claim 1 and asecond compound having the formula:

wherein X is a chain of 0-5 amino acids, inclusive, where the N-terminalamino acid is bonded to R¹ and R² by the nitrogen of the amino group ofthe N-terminal amino acid; Y is a chain of 0-4 amino acids, inclusive,where the C-terminal amino acid has a carboxylamide group, which isindependently bonded to R³ and R⁴; R¹ and R² are each independentlybonded to the amino group of the N-terminal amino acid and selected fromH, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and(C₇-C₁₈)alkaryl; R³ and R⁴ are each independently bonded to the amidegroup of the C-terminus amino acid and selected from H, (C₁-C₁₂)alkyl,(C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; A²²is an aromatic amino acid, Ala, Aib, Anb, N-Me-Ala or is deleted; A²³ isSer, Thr, Ala, Aib, N-Me-Ser, N-Me-Thr, N-Me-Ala or is deleted; A²⁴ isLeu, Ile, Nle, Val, Trp, Gly, Nva, Aib, Anb, N-Me-Leu or is deleted; A²⁵is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-pε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) alkyl group, or an aryl group), Ornor is deleted; A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His,β-pyrazolylalaline, N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg,Lys-ε-NH—R (where R is H, a branched or straight chain (C₁-C₁₀) alkylgroup, an aryl group, or a pharmaceutically acceptable salt thereof),Orn or is deleted; A²⁷ is an aromatic amino acid; A²⁸ is Leu, Ile, Nle,Val, Trp, Aib, Anb or N-Me-Leu; A²⁹ is Asn, Ala, Gln, Gly, Trp orN-Me-Asn; A³⁰ is Leu, Ile, Nle, Nva, Fla, Val, Trp, Aib, Anb orN-Me-Leu; A³¹ is Val, Leu, Nle, Nva, Ile, Trp, Aib, Anb or N-Me-Val; A³²is Thr, Ser, D-Trp, N-Me-Ser or N-Me-Thr; and wherein the compoundoptionally includes one or two pseudopeptide bonds where eachpseudopeptide bond is independently selected from —CH₂—NH—, —CH₂—S—,—CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 10. The method of claim 9 wherein thesecond compound is N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹,ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO. 7) wherein ψ is —CH2-NH—.
 11. Themethod of claim 10 wherein the compound of claim 1 is


12. The method of claim 7 wherein the step of administering to saidsubject the compound of claim 1 includes administering to said subject amixture of the compound of claim 1 and a second compound having theformula:

wherein: the N-terminal amino acid is bonded to R¹ and R²; Y is a chainof 0-4 amino acids, inclusive, where the C-terminal amino acid is bondedto R³ and R⁴ by the side chain of the C-terminal amino acid or by thecarbon of the carboxyl group of the C-terminal amino acid; R¹ and R² areeach independently bonded to the amino group of the N-terminal aminoacid and selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl,(C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; R³ and R⁴ are each independentlybonded to the amide group of the C-terminus amino acid and are selectedfrom H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈)aralkyl, and(C₇-C₁₈)alkaryl; A²⁵ is Arg, Lys, homo-Arg, diethyl-homo-Arg, lys-ε-NH—Rwhere R is H, a branched or straight chain C₁-C₁₀ alkyl group, or anaryl group, Orn or is deleted; A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His,β-pyrozolylalanin, N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg,Lys-e-NH—R where R is H, a branched or straight chain (C₁-C₁₀) alkylgroup, or an aryl group, Orn or is deleted; A²⁷ is an aromatic aminoacid: A²⁸ is Leu, Ile, Val, Trp Nle, Nva, Aib, Anb, or N-Me-Leu; A²⁹ isAsn, Ala, Gin, Fly, Trp, or N-Me-Asn; A³⁰ is Leu, Ile, Val, Trp, Nle,Nva, Aib, Anb, or N-Me-Leu; A³¹ is Val, Ile, Trp, Nva, Aib, Anb, orN-Me-Val; A³² is Thr, Ser, N-Me-Ser, N-Me-Thr, or D-Trp; and wherein thecompound optionally includes one or two pseudopeptide bonds where eachpseudopeptide bond is independently selected from —CH₂—NH—, —CH₂—S—,—CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 13. A therapeutic composition capable ofcontrolling an NPY mediated physiological response comprising atherapeutically effective amount of the compound of claim 1 togetherwith a pharmaceutically acceptable carrier substance.
 14. Thecomposition of claim 13, wherein the therapeutic composition is capableof suppressing appetite.
 15. The composition of claim 13, wherein saidcomposition is in the form of a liquid, pill, tablet, or capsule fororal administration to a subject.
 16. The composition of claim 13,wherein said composition is in the form of a liquid for nasal,intravenous, subcutaneous, parenteral, or intraperitoneal administrationto a subject.
 17. The composition of claim 13 wherein thetherapeutically effective amount of the compound of claim 1 includes atherapeutically effective amount of a mixture of the compound of claim 1and a second compound having the formula:

wherein X is a chain of 0-5 amino acids, inclusive, where the N-terminalamino acid is bonded to R¹ and R² by the nitrogen of the amino group ofthe N-terminal amino acid; Y is a chain of 0-4 amino acids, inclusive,where the C-terminal amino acid has a carboxylamide group, which isindependently bonded to R³ and R⁴; R¹ and R² are each independentlybonded to the amino group of the N-terminal amino acid and selected fromH, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and(C₇-C₁₈)alkaryl; R³ and R⁴ are each independently bonded to the amidegroup of the C-terminus amino acid and selected from H, (C₁-C₁₂)alkyl,(C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈)aralkyl, and (C₇-C₁₈)alkaryl; A²² isan aromatic amino acid, Ala, Aib, Anb, N-Me-Ala or is deleted; A²³ isSer, Thr, Ala, Aib, N-Me-Ser, N-Me-Thr, N-Me-Ala or is deleted; A²⁴ isLeu, Ile, Nle, Val, Trp, Gly, Nva, Aib, Anb, N-Me-Leu or is deleted; A²⁵is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-pε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) alkyl group, or an aryl group), Ornor is deleted; A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His,β-pyrazolylalaline, N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg,Lys-ε-NH—R (where R is H, a branched or straight chain (C₁-C₁₀) alkylgroup, an aryl group, or a pharmaceutically acceptable salt thereof),Orn or is deleted; A²⁷ is an aromatic amino acid; A²⁸ is Leu, Ile, Nle,Val, Trp, Aib, Anb or N-Me-Leu; A²⁹ is Asn, Ala, Gln, Gly, Trp orN-Me-Asn; A³⁰ is Leu, Ile, Nle, Nva, Fla, Val, Trp, Aib, Anb orN-Me-Leu; A³¹ is Val, Leu, Nle, Nva, Ile, Trp, Aib, Anb or N-Me-Val; A³²is Thr, Ser, D-Trp, N-Me-Ser or N-Me-Thr; and wherein the compoundoptionally includes one or two pseudopeptide bonds where eachpseudopeptide bond is independently selected from —CH₂—NH—, —CH₂—S—,—CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 18. The composition of claim 17 whereinthe second compound is N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹,ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO. 7) wherein ψ is —CH2-NH—.
 19. Thecomposition of claim 18 wherein the compound of claim 1 is


20. The composition of claim 13 wherein the therapeutically effectiveamount of the compound of claim 1 includes a therapeutically effectiveamount of a mixture of the compound of claim 1 and a second compoundhaving the formula:

wherein: the N-terminal amino acid is bonded to R¹ and R²; Y is a chainof 0-4 amino acids, inclusive, where the C-terminal amino acid has acarboxylamide group, which is independently bonded to R³ and R⁴; R¹ andR² are each independently bonded to the amino group of the N-terminalamino acid and selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl,(C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; R³ and R⁴ are eachindependently bonded to the amide group of the C-terminus amino acid andare selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl,(C₇-C₁₈)aralkyl, and (C₇-C₁₈)alkaryl; A²⁵ is Arg, Lys, homo-Arg,diethyl-homo-Arg, lys-ε-NH—R where R is H, a branched or straight chainC₁-C₁₀ alkyl group, or an aryl group, Orn or is deleted; A²⁶ is Ala,His, Thr, 3-Me-His, 1-Me-His, β-pyrozolylalanin, N-Me-His, Arg, Lys,homo-Arg, diethyl-homo-Arg, Lys-e-NH—R where R is H, a branched orstraight chain (C₁-C₁₀) alkyl group, or an aryl group, Orn or isdeleted; A²⁷ is an aromatic amino acid: A²⁸ is Leu, Ile, Val, Trp Nle,Nva, Aib, Anb, or N-Me-Leu; A²⁹ is Asn, Ala, Gin, Fly, Trp, or N-Me-Asn;A³⁰ is Leu, Ile, Val, Trp, Nle, Nva, Aib, Anb, or N-Me-Leu; A³¹ is Val,Ile, Trp, Nva, Aib, Anb, or N-Me-Val; A³² is Thr, Ser, N-Me-Ser,N-Me-Thr, or D-Trp; and wherein the compound optionally includes one ortwo pseudopeptide bonds where each pseudopeptide bond is independentlyselected from —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 21. Acompound having the formula:

wherein: each X, independently, is an aromatic amino acid; each Y,independently, is an amino acid having a guanidino group; each Z,independently, is an aliphatic amino acid; A1 and A2, independently, areselected from Cys, Pen, Glu, Asp, Lys, and Dpr; and wherein the compoundoptionally includes one or two pseudopeptide bonds where eachpseudopeptide bond is independently selected from —CH₂—NH—, —CH₂—S—,—CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 22. The compound of claim 21 wherein thecompound includes one or two pseudopeptide bonds where eachpseudopeptide bond is —CH₂—NH—.
 23. The compound of claim 21 wherein thearomatic amino acid is selected from the group consisting of Phe, Trp,and Tyr and the aliphatic amino acid is selected from the groupconsisting of Ile, Leu, Val, Ala, Gly, and Nva.
 24. The compound ofclaim 21 wherein the aromatic amino acid is a hydroxyl aromatic aminoacid.
 25. The compound of claim 21, wherein said compound is


26. A pharmaceutically acceptable salt of the compound of claim
 21. 27.A method for controlling an NPY mediated physiological response in asubject comprising administering to said subject the compound of claim21.
 28. The method of claim 27 wherein the compound is capable ofsuppressing appetite.
 29. The method of claim 27 wherein the step ofadministering to said subject the compound of claim 21 includesadministering to said subject a mixture of the compound of claim 21 anda second compound having the formula:

wherein X is a chain of 0-5 amino acids, inclusive, where the N-terminalamino acid is bonded to R¹ and R² by the nitrogen of the amino group ofthe N-terminal amino acid; Y is a chain of 0-4 amino acids, inclusive,where the C-terminal amino acid has a carboxylamide group, which isindependently bonded to R³ and R⁴; R¹ and R² are each independentlybonded to the amino group of the N-terminal amino acid and selected fromH, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and(C₇-C₁₈)alkaryl; R³ and R⁴ are each independently bonded to the amidegroup of the C-terminus amino acid and selected from H, (C₁-C₁₂)alkyl,(C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; A²²is an aromatic amino acid, Ala, Aib, Anb, N-Me-Ala or is deleted; A²³ isSer, Thr, Ala, Aib, N-Me-Ser, N-Me-Thr, N-Me-Ala or is deleted; A²⁴ isLeu, Ile, Nle, Val, Trp, Gly, Nva, Aib, Anb, N-Me-Leu or is deleted; A²⁵is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-pε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) alkyl group, or an aryl group), Ornor is deleted; A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His,β-pyrazolylalaline, N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg,Lys-ε-NH—R (where R is H, a branched or straight chain (C₁-C₁₀) alkylgroup, an aryl group, or a pharmaceutically acceptable salt thereof),Orn or is deleted; A²⁷ is an aromatic amino acid; A²⁸ is Leu, Ile, Nle,Val, Trp, Aib, Anb or N-Me-Leu; A²⁹ is Asn, Ala, Gln, Gly, Trp orN-Me-Asn; A³⁰ is Leu, Ile, Nle, Nva, Fla, Val, Trp, Aib, Anb orN-Me-Leu; A³¹ is Val, Leu, Nle, Nva, Ile, Trp, Aib, Anb or N-Me-Val; A³²is Thr, Ser, D-Trp, N-Me-Ser or N-Me-Thr; and wherein the compoundoptionally includes one or two pseudopeptide bonds where eachpseudopeptide bond is independently selected from —CH₂—NH—, —CH₂—S—,—CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 30. The method of claim 29 wherein thesecond compound is N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹,ψ^(35/36)]PYY(22-36)-NH₂(SEQ. ID. NO. 7) wherein ψ is —CH2-NH—.
 31. Themethod of claim 30 wherein the compound of claim 21 is


32. The method of claim 27 wherein the step of administering to saidsubject the compound of claim 21 includes administering to said subjecta mixture of the compound of claim 21 and a second compound having theformula:

wherein: the N-terminal amino acid is bonded to R¹ and R²; Y is a chainof 0-4 amino acids, inclusive, where the C-terminal amino acid has acarboxylamide group, which is independently bonded to R³ and R⁴; R¹ andR² are each independently bonded to the amino group of the N-terminalamino acid and selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl,(C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; R³ and R⁴ are eachindependently bonded to the amide group of the C-terminus amino acid andare selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈)aralkyl, and (C₇-C₁₈)alkaryl; A²⁵ is Arg, Lys, homo-Arg,diethyl-homo-Arg, lys-ε-NH—R where R is H, a branched or straight chainC₁-C₁₀ alkyl group, or an aryl group, Orn or is deleted; A²⁶ is Ala,His, Thr, 3-Me-His, 1-Me-His, β-pyrozolylalanin, N-Me-His, Arg, Lys,homo-Arg, diethyl-homo-Arg, Lys-e-NH—R where R is H, a branched orstraight chain (C₁-C₁₀) alkyl group, or an aryl group, Orn or isdeleted; A²⁷ is an aromatic amino acid: A²⁸ is Leu, Ile, Val, Trp Nle,Nva, Aib, Anb, or N-Me-Leu; A²⁹ is Asn, Ala, Gin, Fly, Trp, or N-Me-Asn;A³⁰ is Leu, Ile, Val, Trp, Nle, Nva, Aib, Anb, or N-Me-Leu; A³¹ is Val,Ile, Trp, Nva, Aib, Anb, or N-Me-Val; A³² is Thr, Ser, N-Me-Ser,N-Me-Thr, or D-Trp; and wherein the compound optionally includes one ortwo pseudopeptide bonds where each pseudopeptide bond is independentlyselected from —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 33. Atherapeutic composition capable of controlling an NPY mediatedphysiological response comprising a therapeutically effective amount ofthe compound of claim 21 together with a pharmaceutically acceptablecarrier substance.
 34. The composition of claim 33 wherein thetherapeutic composition is capable of suppressing appetite.
 35. Thecomposition of claim 33 wherein said composition is in the form of aliquid, pill, tablet, or capsule for oral administration to a subject.36. The composition of claim 33 wherein said composition is in the formof a liquid for nasal, intravenous, subcutaneous, parenteral, orintraperitoneal administration to a subject to a subject.
 37. Thecomposition of claim 33 wherein the therapeutically effective amount ofthe compound of claim 21 includes a therapeutically effective amount ofa mixture of the compound of claim 21 and a second compound having theformula:

wherein X is a chain of 0-5 amino acids, inclusive, where the N-terminalamino acid is bonded to R¹ and R² by the nitrogen of the amino group ofthe N-terminal amino acid; Y is a chain of 0-4 amino acids, inclusive,where the C-terminal amino acid has a carboxylamide group, which isindependently bonded to R³ and R⁴; R¹ and R² are each independentlybonded to the amino group of the N-terminal amino acid and selected fromH, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and(C₇-C₁₈)alkaryl; R³ and R⁴ are each independently bonded to the amidegroup of the C-terminus amino acid and selected from H, (C₁-C₁₂)alkyl,(C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; A²²is an aromatic amino acid, Ala, Aib, Anb, N-Me-Ala or is deleted; A²³ isSer, Thr, Ala, Aib, N-Me-Ser, N-Me-Thr, N-Me-Ala or is deleted; A²⁴ isLeu, Ile, Nle, Val, Trp, Gly, Nva, Aib, Anb, N-Me-Leu or is deleted; A²⁵is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-pε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) alkyl group, or an aryl group), Ornor is deleted; A²⁶ is Ala, His, Thr, 3-Me-His, β-Me-His,β-pyrazolylalaline, N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg,Lys-ε—NH—R (where R is H, a branched or straight chain (C₁-C₁₀) alkylgroup, an aryl group, or a pharmaceutically acceptable salt thereof),Orn or is deleted; A²⁷ is an aromatic amino acid; A²⁸ is Leu, Ile, Nle,Val, Trp, Aib, Anb or N-Me-Leu; A²⁹ is Asn, Ala, Gln, Gly, Trp orN-Me-Asn; A³⁰ is Leu, Ile, Nle, Nva, Fla, Val, Trp, Aib, Anb orN-Me-Leu; A³¹ is Val, Leu, Nle, Nva, Ile, Trp, Aib, Anb or N-Me-Val; A³²is Thr, Ser, D-Trp, N-Me-Ser or N-Me-Thr; and wherein the compoundoptionally includes one or two pseudopeptide bonds where eachpseudopeptide bond is independently selected from —CH₂—NH—, —CH₂—S—,—CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 38. The composition of claim 36 whereinthe second compound is N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹,ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO. 7) wherein ψ is —CH2-NH—.
 39. Thecomposition of claim 38 wherein the compound of claim 21 is


40. The composition of claim 33 wherein the therapeutically effectiveamount of the compound of claim 21 includes a therapeutically effectiveamount of a mixture of the compound of claim 21 and a second compoundhaving the formula:

wherein: the N-terminal amino acid is bonded to R¹ and R²; Y is a chainof 0-4 amino acids, inclusive, where the C-terminal amino acid has acarboxylamide group, which is independently bonded to R³ and R⁴; R¹ andR² are each independently bonded to the amino group of the N-terminalamino acid and selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl,(C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; R³ and R⁴ are eachindependently bonded to the amide group of the C-terminus amino acid andare selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈)aralkyl, and (C₇-C₁₈)alkaryl; A²⁵ is Arg, Lys, homo-Arg,diethyl-homo-Arg, lys-ε-NH—R where R is H, a branched or straight chain(C₁-C₁₀) alkyl group, or an aryl group, Orn or is deleted; A²⁶ is Ala,His, Thr, 3-Me-His, 1-Me-His, β-pyrozolylalanin, N-Me-His, Arg, Lys,homo-Arg, diethyl-homo-Arg, Lys-e-NH—R where R is H, a branched orstraight chain (C₁-C₁₀) alkyl group, or an aryl group, Orn or isdeleted; A²⁷ is an aromatic amino acid: A²⁸ is Leu, Ile, Val, Trp Nle,Nva, Aib, Anb, or N-Me-Leu; A²⁹ is Asn, Ala, Gin, Fly, Trp, or N-Me-Asn;A³⁰ is Leu, Ile, Val, Trp, Nle, Nva, Aib, Anb, or N-Me-Leu; A³¹ is Val,Ile, Trp, Nva, Aib, Anb, or N-Me-Val; A³² is Thr, Ser, N-Me-Ser,N-Me-Thr, or D-Trp; and wherein the compound optionally includes one ortwo pseudopeptide bonds where each pseudopeptide bond is independentlyselected from —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 41. Acompound having the formula:H—[X—Y-Z-Y—X]_(n)—NH₂ wherein: each X, independently, is an aromaticamino acid; each Y, independently, is an amino acid having a guanidinogroup; each Z, independently, is an aliphatic amino acid; n=1, 2, 3 or4; and wherein the compound optionally includes one or two pseudopeptidebonds where each pseudopeptide bond is independently selected from—CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 42. The compound ofclaim 41 wherein the compound includes one or two pseudopeptide bondswhere each pseudopeptide bond is —CH₂—NH—.
 43. The compound of claim 41wherein the aromatic amino acid is selected from the group consisting ofPhe, Trp, and Tyr and the aliphatic amino acid is selected from thegroup consisting of Ile, Leu, Val, Ala, Gly, and Nva.
 44. The compoundof claim 41 wherein the aromatic amino acid is a hydroxyl aromatic aminoacid.
 45. The compound of claim 41, wherein said compound is eitherH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ. ID. NO. 9) orH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ.ID. NO. 10).
 46. A pharmaceutically acceptable salt of the compound ofclaim
 41. 47. A method for controlling an NPY mediated physiologicalresponse in a subject comprising administering to said subject thecompound of claim
 41. 48. The method of claim 47 wherein the compound iscapable of suppressing appetite.
 49. The method of claim 47 wherein thestep of administering to said subject the compound of claim 1 includesadministering to said subject a mixture of the compound of claim 1 and asecond compound having the formula:

wherein X is a chain of 0-5 amino acids, inclusive, where the N-terminalamino acid is bonded to R¹ and R² by the nitrogen of the amino group ofthe N-terminal amino acid; Y is a chain of 0-4 amino acids, inclusive,where the C-terminal amino acid has a carboxylamide group, which isindependently bonded to R³ and R⁴; R¹ and R² are each independentlybonded to the amino group of the N-terminal amino acid and selected fromH, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and(C₇-C₁₈)alkaryl; R³ and R⁴ are each independently bonded to the amidegroup of the C-terminus amino acid and selected from H, (C₁-C₁₂)alkyl,(C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; A²²is an aromatic amino acid, Ala, Aib, Anb, N-Me-Ala or is deleted; A²³ isSer, Thr, Ala, Aib, N-Me-Ser, N-Me-Thr, N-Me-Ala or is deleted; A²⁴ isLeu, Ile, Nle, Val, Trp, Gly, Nva, Aib, Anb, N-Me-Leu or is deleted; A²⁵is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-pε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) lkyl group, or an aryl group), Ornor is deleted; A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His,β-pyrazolylalaline, N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg,Lys-ε-NH—R (where R is H, a branched or straight chain (C₁-C₁₀) alkylgroup, an aryl group, or a pharmaceutically acceptable salt thereof),Orn or is deleted; A²⁷ is an aromatic amino acid; A²⁸ is Leu, Ile, Nle,Val, Trp, Aib, Anb or N-Me-Leu; A²⁹ is Asn, Ala, Gln, Gly, Trp orN-Me-Asn; A³⁰ is Leu, Ile, Nle, Nva, Fla, Val, Trp, Aib, Anb orN-Me-Leu; A³¹ is Val, Leu, Nle, Nva, Ile, Trp, Aib, Anb or N-Me-Val; A³²is Thr, Ser, D-Trp, N-Me-Ser or N-Me-Thr; and wherein the compoundoptionally includes one or two pseudopeptide bonds where eachpseudopeptide bond is independently selected from —CH₂—NH—, —CH₂—S—,—CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 50. The method of claim 49 wherein thesecond compound is N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹,ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO. 7) wherein ψ is —CH2-NH—.
 51. Themethod of claim 50 wherein the compound of claim 41 is eitherH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ. ID. NO. 9) orH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ.ID. NO. 10).
 52. The method of claim 47 wherein the step ofadministering to said subject the compound of claim 1 includesadministering to said subject a mixture of the compound of claim 1 and asecond compound having the formula:

wherein: the N-terminal amino acid is bonded to R¹ and R²; Y is a chainof 0-4 amino acids, inclusive, where the C-terminal amino acid has acarboxylamide group, which is independently bonded to R³ and R⁴; R¹ andR² are each independently bonded to the amino group of the N-terminalamino acid and selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl,(C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; R³ and R⁴ are eachindependently bonded to the amide group of the C-terminus amino acid andare selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈)aralkyl, and (C₇-C₁₈)alkaryl; A²⁵ is Arg, Lys, homo-Arg,diethyl-homo-Arg, lys-ε-NH—R where R is H, a branched or straight chain(C₁-C₁₀) alkyl group, or an aryl group, Orn or is deleted; A²⁶ is Ala,His, Thr, 3-Me-His, 1-Me-His, β-pyrozolylalanin, N-Me-His, Arg, Lys,homo-Arg, diethyl-homo-Arg, Lys-e-NH—R where R is H, a branched orstraight chain (C₁-C₁₀) alkyl group, or an aryl group, Orn or isdeleted; A²⁷ is an aromatic amino acid: A²⁸ is Leu, Ile, Val, Trp Nle,Nva, Aib, Anb, or N-Me-Leu; A²⁹ is Asn, Ala, Gin, Fly, Trp, or N-Me-Asn;A³⁰ is Leu, Ile, Val, Trp, Nle, Nva, Aib, Anb, or N-Me-Leu; A³¹ is Val,Ile, Trp, Nva, Aib, Anb, or N-Me-Val; A³² is Thr, Ser, N-Me-Ser,N-Me-Thr, or D-Trp; and wherein the compound optionally includes one ortwo pseudopeptide bonds where each pseudopeptide bond is independentlyselected from —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 53. Atherapeutic composition capable of controlling an NPY mediatedphysiological response comprising a therapeutically effective amount ofthe compound of claim 41 together with a pharmaceutically acceptablecarrier substance.
 54. The composition of claim 53 wherein thetherapeutic composition is capable of suppressing appetite.
 55. Thecomposition of claim 53 wherein said composition is in the form of aliquid, pill, tablet, or capsule for oral administration to a subject.56. The composition of claim 53 wherein said composition is in the formof a liquid for nasal, intravenous, subcutaneous, parenteral, orintraperitoneal administration to a subject to a subject.
 57. Thecomposition of claim 53 wherein the therapeutically effective amount ofthe compound of claim 41 includes a therapeutically effective amount ofa mixture of the compound of claim and a second compound having theformula:

wherein X is a chain of 0-5 amino acids, inclusive, where the N-terminalamino acid is bonded to R¹ and R² by the nitrogen of the amino group ofthe N-terminal amino acid; Y is a chain of 0-4 amino acids, inclusive,where the C-terminal amino acid has a carboxylamide group, which isindependently bonded to R³ and R⁴; R¹ and R² are each independentlybonded to the amino group of the N-terminal amino acid and selected fromH, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and(C₇-C₁₈)alkaryl; R³ and R⁴ are each independently bonded to the amidegroup of the C-terminus amino acid and selected from H, (C₁-C₁₂)alkyl,(C₆-C₁₈)aryl, (C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; A²²is an aromatic amino acid, Ala, Aib, Anb, N-Me-Ala or is deleted; A²³ isSer, Thr, Ala, Aib, N-Me-Ser, N-Me-Thr, N-Me-Ala or is deleted; A²⁴ isLeu, Ile, Nle, Val, Trp, Gly, Nva, Aib, Anb, N-Me-Leu or is deleted; A²⁵is Arg, Lys, homo-Arg, diethyl-homo-Arg, Lys-pε-NH—R (where R is H, abranched or straight chain (C₁-C₁₀) alkyl group, or an aryl group), Ornor is deleted; A²⁶ is Ala, His, Thr, 3-Me-His, 1-Me-His,β-pyrazolylalaline, N-Me-His, Arg, Lys, homo-Arg, diethyl-homo-Arg,Lys-ε-NH—R (where R is H, a branched or straight chain (C₁-C₁₀) alkylgroup, an aryl group, or a pharmaceutically acceptable salt thereof),Orn or is deleted; A²⁷ is an aromatic amino acid; A²⁸ is Leu, Ile, Nle,Val, Trp, Aib, Anb or N-Me-Leu; A²⁹ is Asn, Ala, Gln, Gly, Trp orN-Me-Asn; A³⁰ is Leu, Ile, Nle, Nva, Fla, Val, Trp, Aib, Anb orN-Me-Leu; A³¹ is Val, Leu, Nle, Nva, Ile, Trp, Aib, Anb or N-Me-Val; A³²is Thr, Ser, D-Trp, N-Me-Ser or N-Me-Thr; and wherein the compoundoptionally includes one or two pseudopeptide bonds where eachpseudopeptide bond is independently selected from —CH₂—NH—, —CH₂—S—,—CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 58. The composition of claim 57 whereinthe second compound is N-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹,ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO. 7) wherein ψ is —CH2-NH—.
 59. Thecomposition of claim 58 wherein the compound of claim 41 is eitherH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ. ID. NO. 9) orH-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-Tyr-Arg-Leu-Arg-Tyr-NH₂ (SEQ.ID. NO. 10).
 60. The composition of claim 53 wherein the therapeuticallyeffective amount of the compound of claim 41 includes a therapeuticallyeffective amount of a mixture of the compound of claim and a secondcompound having the formula:

wherein: the N-terminal amino acid is bonded to R¹ and R²; Y is a chainof 0-4 amino acids, inclusive, where the C-terminal amino acid has acarboxylamide group, which is independently bonded to R³ and R⁴; R¹ andR² are each independently bonded to the amino group of the N-terminalamino acid and selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl,(C₁-C₁₂)acyl, (C₇-C₁₈) aralkyl, and (C₇-C₁₈)alkaryl; R³ and R⁴ are eachindependently bonded to the amide group of the C-terminus amino acid andare selected from H, (C₁-C₁₂)alkyl, (C₆-C₁₈)aryl, (C₁-C₁₂)acyl,(C₇-C₁₈)aralkyl, and (C₇-C₁₈)alkaryl; A²⁵ is Arg, Lys, homo-Arg,diethyl-homo-Arg, lys-ε-NH—R where R is H, a branched or straight chain(C₁-C₁₀) alkyl group, or an aryl group, Orn or is deleted; A²⁶ is Ala,His, Thr, 3-Me-His, 1-Me-His, β-pyrozolylalanin, N-Me-His, Arg, Lys,homo-Arg, diethyl-homo-Arg, Lys-e-NH—R where R is H, a branched orstraight chain (C₁-C₁₀) alkyl group, or an aryl group, Orn or isdeleted; A²⁷ is an aromatic amino acid: A²⁸ is Leu, Ile, Val, Trp Nle,Nva, Aib, Anb, or N-Me-Leu; A²⁹ is Asn, Ala, Gin, Fly, Trp, or N-Me-Asn;A³⁰ is Leu, Ile, Val, Trp, Nle, Nva, Aib, Anb, or N-Me-Leu; A³¹ is Val,Ile, Trp, Nva, Aib, Anb, or N-Me-Val; A³² is Thr, Ser, N-Me-Ser,N-Me-Thr, or D-Trp; and wherein the compound optionally includes one ortwo pseudopeptide bonds where each pseudopeptide bond is independentlyselected from —CH₂—NH—, —CH₂—S—, —CH₂—CH₂—, —CH₂—O— and CH₂—CO—.
 61. Amethod for controlling an NPY mediated physiological response in asubject comprising administering to said subject a mixture of NPY Y₄ andY₂ selective receptor agonists.
 62. The method of claim 61 wherein theY₄ receptor agonist is PP, a peptide fragment thereof, or an analog ofPP or the peptide fragment, and the Y₂ receptor agonist is PYY(3-36), apeptide fragment thereof, or an analog of PYY(3-36) or the peptidefragment.
 63. The method of claim 62 wherein the Y₄ receptor agonist isthe peptide fragment PP(32-36) or an analog thereof and the Y₂ receptoragonist is the peptide fragment PYY(22-36), or an analog thereof. 64.The method of claim 61, wherein the mixture is capable of suppressingappetite.
 65. A method for controlling an NPY mediated physiologicalresponse in a subject comprising administering to said subject, togetherwith a pharmaceutically acceptable carrier substance, a therapeuticcomposition comprising a therapeutically effective amount of a mixtureof NPY Y₂ and Y₄ receptor agonists.
 66. The method of claim 65 whereinthe Y₄ receptor agonist is PP, a peptide fragment thereof, or an analogof PP or the peptide fragment, and the Y₂ receptor agonist is PYY(3-36),a peptide fragment thereof, or an analog of PYY(3-36) or the peptidefragment.
 67. The method of claim 66 wherein the Y₄ receptor agonist isthe peptide fragment PP(32-36) or an analog thereof and the Y₂ receptoragonist is the peptide fragment PYY(22-36), or an analog thereof. 68.The method of claim 67 wherein the analog of PYY(22-36) isN-α-Ac[Nle^(24,28), Trp³⁰, Nva³¹, ψ^(35/36)]PYY(22-36)-NH₂ (SEQ. ID. NO.7) wherein ψ is —CH2-NH— and the analog of PP(32-36) is either


69. The method of claim 65, wherein the mixture is capable ofsuppressing appetite.
 70. The method of claim 65 wherein saidcomposition is in the form of a liquid, pill, tablet, or capsule fororal administration to a subject.
 71. The method of claim 65 whereinsaid composition is in the form of a liquid for nasal, intravenous,subcutaneous, parenteral, or intraperitoneal administration to asubject.